U.S. patent application number 13/787534 was filed with the patent office on 2013-09-26 for electrochemical sensors.
The applicant listed for this patent is Sensor Innovations, Inc.. Invention is credited to Carolyn R. Kahn, Dean Mansour, Vern Norviel, Robbie Singh Sidhu, James A. Wilkins, Elicia Wong.
Application Number | 20130248364 13/787534 |
Document ID | / |
Family ID | 46245407 |
Filed Date | 2013-09-26 |
United States Patent
Application |
20130248364 |
Kind Code |
A1 |
Kahn; Carolyn R. ; et
al. |
September 26, 2013 |
ELECTROCHEMICAL SENSORS
Abstract
Systems and methods are provided for detecting the presence of
an analyte in a sample. A solid state electrochemical sensor can
include a redox active moiety having an oxidation and/or reduction
potential that is sensitive to the presence of an analyte
immobilized over a surface of a working electrode. A redox active
moiety having an oxidation and/or reduction potential that is
insensitive to the presence of the analyte can be used for
reference. Voltammetric measurements made using such systems can
accurately determine the presence and/or concentration of the
analyte in the sample. The solid state electrochemical sensor can
be robust and not require calibration or re-calibration.
Inventors: |
Kahn; Carolyn R.; (San
Francisco, CA) ; Wong; Elicia; (Foster City, CA)
; Wilkins; James A.; (San Francisco, CA) ;
Mansour; Dean; (Redwood City, CA) ; Norviel;
Vern; (San Francisco, CA) ; Sidhu; Robbie Singh;
(Fremont, CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Sensor Innovations, Inc. |
Burlingame |
CA |
US |
|
|
Family ID: |
46245407 |
Appl. No.: |
13/787534 |
Filed: |
March 6, 2013 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13329135 |
Dec 16, 2011 |
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13787534 |
|
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61424040 |
Dec 16, 2010 |
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61550355 |
Oct 21, 2011 |
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Current U.S.
Class: |
204/433 |
Current CPC
Class: |
Y10T 428/24421 20150115;
Y02A 90/26 20180101; G01N 27/3335 20130101; Y02A 90/10 20180101;
G01N 27/4167 20130101; G01N 27/302 20130101 |
Class at
Publication: |
204/433 |
International
Class: |
G01N 27/30 20060101
G01N027/30 |
Claims
1. A sensor for detecting the presence or absence of an analyte,
comprising: an electrode having a surface having immobilized
thereon a redox-active moiety, wherein the redox-active moiety
exhibits an oxidation potential and/or a reduction potential that
is sensitive or insensitive to the presence of the analyte; and a
light-emitting device adjacent to said electrode, said
light-emitting device configured to generate light.
2. The sensor of claim 1, wherein the light-emitting device is
configured to generate light that is i) incident on said surface,
ii) incident on another surface of said electrode, said another
surface opposite from said surface, and/or iii) directed through
said electrode.
3. The sensor of claim 1, wherein the electrode is formed of a
semiconductor material.
4. The sensor of claim 3, wherein the semiconductor material
includes silicon.
5. The sensor of claim 1, wherein the electrode is formed of a
non-semiconductor material.
6. The sensor of claim 5, wherein the non-semiconductor material
includes carbon.
7. The sensor of claim 1, wherein the light-emitting device is a
light-emitting diode having an active region configured to generate
light upon the recombination of electrons and holes.
8. The sensor of claim 7, wherein the light-emitting diode is an
organic light-emitting diode.
9. The sensor of claim 1, wherein light from said light-emitting
device is incident on said surface.
10. A sensor for detecting the presence or absence of an analyte,
comprising a working electrode having a redox active moiety formed
adjacent to a light emitting device.
11. The sensor of claim 10, wherein the working electrode is formed
of a semiconductor material.
12. The sensor of claim 11, wherein the semiconductor material
comprises silicon.
13. The sensor of claim 10, wherein the working electrode is formed
of a non-semiconductor material.
14. The sensor of claim 13, wherein the non-semiconductor material
comprises carbon.
15. The sensor of claim 10, wherein the light-emitting device is a
light-emitting diode having an active region configured to generate
light upon the recombination of electrons and holes.
16. A solid state sensor for detecting the presence or absence of
an analyte, comprising a solid state electrode configured to detect
the presence or absence of the analyte, and a light emitting device
adjacent to the solid state electrode.
17. The solid state sensor of claim 16, wherein the solid state
electrode is formed of a semiconductor material.
18. The solid state sensor of claim 17, wherein the semiconductor
material comprises silicon.
19. The solid state sensor of claim 16, wherein the solid state
electrode is formed of a non-semiconductor material.
20. The solid state sensor of claim 19, wherein the
non-semiconductor material comprises carbon.
Description
CROSS-REFERENCE
[0001] This application is a continuation of U.S. patent
application Ser. No. 13/329,135 filed Dec. 16, 2011 which claims
the benefit the benefit under 35 USC 119(e) of U.S. Provisional
Patent Application Ser. No. 61/424,040, filed Dec. 16, 2010, and
U.S. Provisional Patent Application Ser. No. 61/550,355, filed Oct.
21, 2011, which are entirely incorporated herein by reference.
BACKGROUND OF THE INVENTION
[0002] The measurement of analyte concentration, in particular,
hydrogen ion concentration or pH is important in a number of
research, industrial, and manufacturing processes. For instance,
the measurement of pH is routinely practiced in food and beverage,
biofuel, biophamaceuticals, as well as in the treatment of water
and waste.
[0003] Many conventional pH sensors use a potentiometric approach
which involves the use of glass electrode to measure pH. The
potentiometric approach suffers from several drawbacks. One
limitation of potentiometric sensors is the need for constant
calibration. Potentiometric pH electrodes, like batteries, tend to
run down with time and use. As the potentiometric electrode ages,
its glass membrane tends to change in resistance, which in turn
will alter the electrode potential. For this reason, the glass
electrodes require calibration on a regular basis. The need for
constant recalibration to provide an accurate pH output
significantly impedes industrial applications especially where
constant in-line pH measurements are required. Recalibration is
particularly cumbersome in a biotech environment where pH
measurement is conducted in medium containing biological materials.
Another significant drawback of conventional pH sensors is that the
glass electrodes have internal solutions, which in some cases can
leak out into the solution being measured. The glass electrodes can
also become fouled by species in the measuring solution, e.g.,
proteins, causing the glass electrode to foul. ISFET devices have
been developed which use a field effect transistor structure on a
silicon surface to measure pH (Bergveld Em et al., IEEE Sensor
Conference, Toronto, October 2003, 1-26). These devices also have
limitations. Thus, there remains a considerable need for reliable
and consistent analyte sensors, and in particular, pH sensors.
SUMMARY OF THE INVENTION
[0004] An aspect of the invention provides a sensor for detecting
the presence of an analyte comprising: a semiconductor electrode
having a surface having immobilized thereon a redox-active moiety,
wherein the redox-active moiety exhibits an oxidation potential
and/or a reduction potential that is sensitive to the presence of
the analyte. In some embodiments the analyte is hydrogen ion and
the redox-active moiety is sensitive to hydrogen ion concentration.
In some embodiments the sensor comprises a plurality of
redox-active moieties, wherein at least one of the redox-active
moieties is sensitive to the presence of an analyte, and at least
one other redox-active moiety is substantially insensitive to the
presence of the analyte.
[0005] In some embodiments the analyte is hydrogen ion, and the
moiety that is substantially insensitive to the presence of
hydrogen ion has a substituent selected from the group consisting
of ferrocene, polyvinylferrocene, viologen, polyviologen, and
polythiophene. In some embodiments the moiety that is substantially
insensitive to the presence of hydrogen ion is ferrocene or a
derivative of ferrocene.
[0006] In some embodiments the analyte is hydrogen, and the
redox-active moiety that is sensitive to the presence of the
hydrogen ion comprises a substituent selected from the group
consisting of anthracenes, quinones, anthroquinones,
phenanthroquinones, phenylene diamines, catechols, phenothiazinium
dyes, monoquaternized N-alkyl-4,4'-bipyridinium, RuOx, and Ni(OH)2.
In some embodiments the redox-active moiety that is sensitive to
the presence of hydrogen ion comprises a substituent comprising
anthracene. In some embodiments the redox-active moiety that is
sensitive to the presence of hydrogen ion comprises a substituent
comprising an anthraquinone or a phenanthraquinone.
[0007] In some embodiments the redox-active moiety that is
sensitive to the presence of an analyte is sensitive to the
concentration of the analyte. In some embodiments the oxidation
potential and/or reduction potential of the redox-active moiety is
sensitive to the concentration of the analyte in a range from
10.sup.-3 M to 10.sup.-10 M. In some embodiments oxidation
potential and/or reduction potential of the redox-active moiety is
sensitive to the concentration of the analyte in a range from
10.sup.-1 M to 10.sup.-14 M. In some embodiments the sensor detects
hydrogen ion concentration from pH 3 to pH 10. In some embodiments
the sensor detects hydrogen ion concentration from pH 1 to pH
14.
[0008] In some embodiments the sensor measures hydrogen ion
concentration within an accuracy of plus or minus 0.1 pH unit. In
some embodiments the sensor measures hydrogen ion concentration
within an accuracy of plus or minus 0.01 pH units.
[0009] In some embodiments the redox-active moiety is covalently
bound to the surface of the electrode. In some embodiments the
redox-active moiety is bound to a polymer that is immobilized onto
the surface of the electrode.
[0010] In some embodiments the semiconductor electrode is doped. In
some embodiments the semiconductor electrode is P-doped. In some
embodiments the semiconductor electrode comprises a silicon
electrode doped with boron. In some embodiments the semiconductor
electrode is N-doped. In some embodiments the semiconductor
electrode comprises a silicon electrode is doped with
phosphorous.
[0011] In some embodiments the semiconductor electrode comprises a
monolithic piece of silicon.
[0012] In some embodiments the semiconductor electrode comprises a
composite electrode, the composite electrode comprising
semiconductor particles in a matrix. In some embodiments the
semiconductor electrode comprises a composite electrode bound to a
conductive substrate. In some embodiments the semiconductor
particles are present in an amount such that a conductive path is
created across the composite electrode.
[0013] In some embodiments the electrode comprises silicon. In some
embodiments the electrode comprises unpolished silicon. In some
embodiments the electrode comprises polished silicon.
[0014] In some embodiments the semiconductor electrode has a
resistivity within the range of 0.01 to 1000 .OMEGA.-cm. In some
embodiments the semiconductor electrode has a resistivity within
the range of 1 to 100 .OMEGA.-cm.
[0015] In some embodiments the sensor is capable of measuring
analyte concentration without calibration with an external
standard. In some embodiments the sensor remains sensitive to the
analyte without calibration after a first use by an end user. In
some embodiments the analyte is hydrogen ion and the sensor remains
sensitive to hydrogen ion after exposure to a cell culture medium
for at least 6 days. In some embodiments the sensor is capable of
measuring pH with an accuracy of plus or minus 0.2 pH units after
exposure to the cell culture medium for at least 6 days. In some
embodiments the analyte is hydrogen ion and the sensor is capable
of measuring pH with an accuracy of plus or minus 0.2 pH units
after autoclave treatment at 121.degree. C. for 40 minutes. In some
embodiments the sensor is capable of measuring pH with an accuracy
of plus or minus 0.2 units after autoclave treatment at 121.degree.
C. for 400 minutes.
[0016] In some embodiments the semiconductor substrate has a
plurality of zones wherein at least a first zone is sensitive to an
analyte, and a second zone that is insensitive to an analyte.
[0017] Another aspect of the invention provides an analyte-sensing
system comprising: a working electrode having a semiconductor
surface that has immobilized thereon a redox-active moiety, wherein
the redox-active moiety has an oxidation potential and/or reduction
potential that is sensitive to the presence of the analyte; a
counter electrode and optionally a reference electrode; a source
for supplying a plurality of potentials to the working electrode;
and a device for measuring current through the working electrode at
the plurality of potentials.
[0018] In some embodiments the invention further comprises a second
working electrode comprising a second semiconductor substrate
comprising a second redox-active moiety having an oxidation
potential and/or reduction potential that is insensitive to the
presence of the analyte. In some embodiments the source for
supplying a plurality of potentials is a potentiostat capable of
applying square waves for square wave voltammetry.
[0019] In some embodiments the invention further comprises a
computation system that communicates with the device for measuring
current, and that calculates a reduction or oxidation potential
from the measured current at a plurality of potentials.
[0020] In some embodiments the system is used as an in-line sensor
in a process.
[0021] In some embodiments the currents measured at a plurality of
potentials are used to determine analyte concentration, and the
determined analyte concentration is used to control a process
parameter.
[0022] In some embodiments the sensor measures hydrogen ion
concentration within an accuracy of plus or minus 0.1 pH unit. In
some embodiments the sensor measures hydrogen ion concentration
within an accuracy of plus or minus 0.01 pH units.
[0023] In some embodiments the sensor is capable of measuring
analyte concentration without calibration with an external
standard. In some embodiments the sensor remains sensitive to the
analyte without calibration after a first use by an end user. In
some embodiments the analyte is hydrogen ion and the sensor remains
sensitive to hydrogen ion after exposure to a cell culture medium
for at least 6 days. In some embodiments the sensor is capable of
measuring pH with an accuracy of plus or minus 0.2 units after
exposure to the cell culture medium for at least 6 days. In some
embodiments the analyte is hydrogen ion and the sensor is capable
of measuring pH with an accuracy of plus or minus 0.2 units after
autoclave treatment at 121.degree. C. for 40 minutes. In some
embodiments the sensor is capable of measuring pH with an accuracy
of plus or minus 0.2 units after autoclave treatment at 121.degree.
C. for 400 minutes.
[0024] Another aspect of the invention provides a semiconductor
substrate having a surface, wherein the surface comprises a
redox-active moiety immobilized thereon, the redox-active moiety
exhibiting an oxidation potential and/or a reduction potential that
is sensitive to the presence of an analyte.
[0025] In some embodiments immobilized thereon is also a second
redox-active moiety having an oxidation potential and/or a
reduction potential that is substantially insensitive to the
presence of the analyte.
[0026] In some embodiments the analyte is an ion. In some
embodiments the analyte is hydrogen ion. In other embodiments, the
analyte is a polarizable molecule. In other embodiments, the
analyte is an ionizable molecule, such as upon the application of
an external source of energy.
[0027] In some embodiments the semiconductor comprises an inorganic
semiconductor. In some embodiments the semiconductor comprises an
organic semiconductor. In some embodiments the inorganic
semiconductor comprises silicon or gallium arsenide. In some
embodiments the organic semiconductor comprises polyacetylene,
polythiophene, or polypyrrole.
[0028] In some embodiments the semiconductor comprises silicon. In
some embodiments the semiconductor comprises unpolished silicon. In
some embodiments the semiconductor comprises polished silicon.
[0029] In some embodiments the redox-active moiety is directly
bound to the surface. In some embodiments the redox-active moiety
is covalently bound to the surface. In some embodiments the
redox-active moiety is covalently bound to the surface through a
linker. In some embodiments the redox-active moiety is covalently
bound to a polymer that is immobilized onto the surface of the
semiconductor substrate. In some embodiments the redox-active
moiety is covalently bound to a polymer that is covalently bound to
the surface of the semiconductor substrate
[0030] In some embodiments the semiconductor is doped. In some
embodiments the semiconductor is N-doped. In some embodiments the
semiconductor is P-doped.
[0031] In some embodiments substrate comprises crystalline silicon
wherein the surface displaying predominantly one crystalline
plane.
[0032] In some embodiments the substrate has a plurality of zones
wherein each zone comprises a redox-active moiety immobilized
thereon, the redox-active moiety exhibiting an oxidation potential
and/or a reduction potential that is sensitive to the presence of
an analyte. In some embodiments a first zone comprises a redox
moiety sensitive to a first analyte, and a second zone comprises a
redox moiety sensitive to a second analyte. In some embodiments the
invention further comprises a third redox-active moiety exhibiting
an oxidation potential and/or a reduction potential that is
sensitive to the presence of a second analyte. In some embodiments
the second analyte is ammonia, oxygen or carbon dioxide.
[0033] Another aspect of the invention provides a method for
forming an analyte-sensitive semiconductor electrode, the electrode
having a surface, the method comprising immobilizing a redox-active
moiety that is sensitive to the presence of an analyte onto the
surface. In some embodiments immobilizing the redox-active moiety
covalently binds the redox-active moiety to the surface. In some
embodiments the redox-active moiety is covalently bound to the
surface through a linker. In some embodiments immobilizing the
redox-active moiety comprises hydrosilation, a free radical
reaction, carbodiimide coupling, a Diels-Alder reaction, a Michael
addition, or click chemistry.
[0034] In some embodiments the redox-active moiety is covalently
bound to a polymer that is immobilized onto the surface.
[0035] In some embodiments the step of immobilizing comprises
polymerization including a monomer or oligomer comprising a
redox-active moiety. In some embodiments the polymerization of a
monomer or oligomer comprising a redox-active moiety includes a
reaction with a functional group covalently bound to the surface,
whereby the polymer formed by polymerization is covalently bound to
the surface. In some embodiments the monomer or oligomer is
electropolymerized onto the surface.
[0036] In some embodiments the step of immobilizing comprises
coating or casting the polymer onto the surface.
[0037] In some embodiments the semiconductor surface comprises a
composite electrode that comprises semiconductor particles within a
matrix.
[0038] Another aspect of the invention provides a method for
forming a semiconductor surface derivatized with one or more
redox-active moieties comprising: contacting an H-terminated
semiconductor surface with the one or more redox-active moieties
wherein at least one redox active moiety is sensitive to the
presence of an analyte, and wherein each redox-active moiety
comprises a functional group that will react with the H-terminated
semiconductor surface to form a covalently bond, thereby forming a
derivatized semiconductor surface.
[0039] In some embodiments the semiconductor surface comprises
silicon.
[0040] In some embodiments at least two redox active moieties are
used, and one of the redox active moieties is insensitive to the
presence of the analyte.
[0041] In some embodiments the H-terminated semiconductor surface
is formed by treatment with hydrofluoric acid.
[0042] In some embodiments the functional group is a vinyl group.
In some embodiments the functional group is an aldehyde group. In
some embodiments the functional group is a diazonium group.
[0043] Another aspect of the invention provides a method of
determining the concentration of an analyte, comprising placing an
electrode in contact with the analyte, the electrode comprising a
semiconductor substrate with a semiconductor surface having
immobilized thereon an analyte-sensitive redox-active moiety, the
analyte-sensitive redox-active moiety exhibiting an oxidation
potential and/or reduction potential that is sensitive to the
concentration of the analyte; applying a plurality of potentials to
the electrode; and measuring the current through the electrode at
the plurality of potentials to determine a reduction and/or
oxidation potential of the analyte-sensitive redox-active moiety,
thereby determining the concentration of the analyte.
[0044] In some embodiments the measuring of the current at the
plurality of potentials provides a peak current, and whereby the
peak current is used to determine reduction and/or oxidation
potential of the analyte-sensitive redox-active moiety.
[0045] In some embodiments the analyte is hydrogen ion.
[0046] In some embodiments the invention further comprises an
analyte-insensitive redox-active moiety bound to a an electrode
comprising a semiconductor substrate, such redox-active moiety
having a reduction and/or oxidation potential that is substantially
insensitive to the analyte, further comprising determining the
oxidation and/or reduction potential of the analyte-insensitive
redox-active moiety, and determining the concentration of the
analyte from the difference in the oxidation and/or reduction
potentials of the analyte-sensitive and analyte-insensitive
moieties.
[0047] In some embodiments the analyte is provided in a
solution.
[0048] Another aspect of the invention provides a sensor for
detecting the presence of an analyte. The sensor comprises a
semiconductor electrode having a surface having immobilized thereon
a layer of a redox-active moiety, wherein the redox-active moiety
exhibits an oxidation potential and/or a reduction potential that
is sensitive or insensitive to the presence of the analyte, and a
layer of composite material on, over or adjacent to the layer of
the redox-active moiety. In some cases, the layer of the composite
material covers the layer of the redox-active moiety.
[0049] In some embodiments, the composite material comprises
Nafion. In some cases, the composite material comprises a porous
material, such as a porous polymeric material (e.g., plastic),
impregnated with Nafion.
[0050] In some situations, a working electrode comprises a layer of
a polymeric material for shielding light-sensitive moieties on or
over the working electrode from light. In some cases, the layer of
the polymeric material comprises polyethersulphone (PES).
[0051] In another aspect of the invention, an electrochemical
sensor comprises a solid state (e.g., a semiconductor, such as
silicon, or carbon) electrode that is equipped with (or operatively
coupled to) a light emitting device (also "light source"
herein).
[0052] In some embodiments, a sensor comprises a light-emitting
device, such as a light-emitting diode, and a layer of a
semiconducting or a non-semiconducting material over the light
emitting device. The layer of the semiconducting or
non-semiconducting material may have a redox sensitive moiety
thereon.
[0053] In some embodiments, the light-emitting diode is an organic
light-emitting diode. In some embodiments, the semiconducting
material is silicon. In some embodiments, the non-semiconducting
material is carbon, such as activated carbon.
[0054] In another embodiment, a sensor for detecting the presence
or absence of an analyte comprises an electrode having a surface
having immobilized thereon a redox-active moiety, wherein the
redox-active moiety exhibits an oxidation potential and/or a
reduction potential that is sensitive or insensitive to the
presence of the analyte. A light-emitting device is adjacent to the
electrode. The light-emitting device is configured to generate
light.
[0055] In some embodiments, a solid state sensor for detecting the
presence or absence of an analyte, comprises a solid state
electrode configured to detect the presence or absence of the
analyte, and a light emitting device adjacent to the solid state
electrode.
[0056] In some embodiments, a sensor for detecting the presence or
absence of an analyte, comprising a working electrode having a
redox active moiety formed adjacent a light emitting device, is
provided.
[0057] In another aspect of the invention, a sensor for detecting
the presence or absence of an analyte comprises a semiconductor
electrode having a surface having immobilized thereon a layer of a
redox-active moiety, wherein the redox-active moiety exhibits an
oxidation potential and/or a reduction potential that is sensitive
or insensitive to the presence of the analyte. A light blocking
layer is adjacent to the layer of the redox-active moiety.
[0058] In some embodiments, a solid state sensor for detecting the
presence or absence of an analyte, comprises a solid state
electrode and a light blocking layer adjacent to the solid state
electrode. The light blocking layer may be formed of a polymeric
material. In some situations, the light blocking layer transmits
less than 10%, 5%, or 1% of light incident on the light blocking
layer.
[0059] Another aspect of the invention is a method comprising:
measuring a pH value of a step in a water or waste treatment
process with a voltammetric pH sensor, wherein the pH sensor
comprises a redox-active moiety that is sensitive to hydrogen ion
concentration, and a redox-active moiety that is substantially
insensitive to hydrogen ion; and using the pH value to monitor or
control the treatment process.
[0060] Another aspect of the invention provides a method comprising
measuring a pH value of a reaction mixture in a biopharmaceutical
process with a voltammetric pH sensor, wherein the pH sensor
comprises a redox-active moiety that is sensitive to hydrogen ion
concentration, and a redox-active moiety that is substantially
insensitive to hydrogen ion concentration to obtain a pH value. The
pH value is used to monitor the biopharmaceutical process.
[0061] In some embodiments the pH value is measured on a sample
obtained from the reaction mixture.
[0062] Another aspect of the invention provides a reactor for
carrying out a biopharmaceutical process wherein the reactor
comprises a pH sensor having a redox-active moiety that is
sensitive to hydrogen ion concentration, and a redox-active moiety
that is substantially insensitive to hydrogen ion
concentration.
[0063] In some embodiments the pH sensor is a voltammetric pH
sensor.
[0064] In an embodiment, the pH sensor is a disposable pH sensor.
In another embodiment, the pH sensor is a single-use pH sensor. In
another embodiment, the pH sensor is a disposable and single-use pH
sensor.
[0065] In some embodiments the reactor is a disposable bioreactor.
In some embodiments the reactor is a bioprocess flexible
container.
[0066] Another aspect of the invention provides a method for
carrying out an industrial process comprising: measuring a pH value
of a step of an industrial process with a voltammetric pH sensor
having a redox-active moiety that is sensitive to hydrogen ion
concentration, and a redox-active moiety that is substantially
insensitive to hydrogen ion concentration; and using the pH value
to carry out the industrial process.
[0067] Another aspect of the invention provides a sensor for
measuring ion concentration in a bodily fluid within a body. The
sensor comprises an electrode configured to be in contact with a
bodily fluid, the electrode comprising a semiconductor surface that
has immobilized thereon a redox-active moiety. The redox-active
moiety has an oxidation potential and/or reduction potential that
is sensitive to concentration of the ion.
[0068] Another aspect of the invention provides a method for
measuring concentration in a bodily fluid within a body, comprising
placing such a sensor in contact with the bodily fluid and
operating the sensor to yield a value of the concentration of the
ion present in the bodily fluid.
[0069] Another aspect of the invention provides a bioreactor
comprising a reservoir for containing a reaction mixture and a pH
probe wherein the pH probe comprises an electrode having a
semiconductor surface, the semiconductor surface having immobilized
thereon a redox active moiety having a reduction and/or oxidation
potential that is sensitive to the presence of hydrogen ion.
[0070] In some embodiments the invention further comprises a
semiconductor surface having immobilized thereon a redox active
moiety having a reduction and/or oxidation potential that is
insensitive to the presence of hydrogen ion.
[0071] In some embodiments the semiconductor surface on which the
redox active moiety having a reduction and/or oxidation potential
that is sensitive to the presence of hydrogen ion is immobilized is
the same semiconductor surface on which the redox active moiety
having a reduction and/or oxidation potential that is insensitive
to the presence of hydrogen ion is immobilized on. In some
embodiments the probe further comprises a counter electrode.
[0072] Another aspect of the invention provides a sensor system,
comprising a redox-active moiety-containing analyte sensor for
insertion into a container for use with a glass probe analyte
sensor. In an embodiment, the redox-active moiety-containing
analyte sensor comprises one or more redox-active moieties. In
another embodiment, the redox-active moiety-containing analyte
sensor comprises a redox-active moiety that is sensitive to the
presence of an analyte and another redox-active moiety that is
insensitive to the presence of the analyte. In another embodiment,
the redox-active moiety-containing analyte sensor is disposed in a
probe body having a form factor configured for insertion into a
container for use with a glass probe analyte sensor.
[0073] Another aspect of the invention provides a method for
detecting the presence or absence of an analyte, comprising using a
sensor system, as described above, to detect the presence or
absence of the analyte.
[0074] Another aspect of the invention provides a method for
forming an analyte sensor, comprising inserting a sensor system as
described herein into a container configured for use with a glass
probe analyte sensor. In an embodiment, the method further
comprises removing a glass probe analyte sensor from the container
prior to inserting the sensor system into the container.
[0075] Another aspect of the invention provides a method for
forming an analyte sensor, comprising inserting a sensor system as
described herein into a container configured for use with a
reactor, in-line flow system, or sample preparation, or
analysis.
[0076] Another aspect of the invention provides a sensor for
detecting the presence or absence of an analyte, comprising a
semiconductor electrode having a surface having immobilized thereon
a redox-active moiety, wherein the redox-active moiety exhibits an
oxidation potential and/or a reduction potential that is sensitive
to the presence of the analyte. The sensor includes a form factor
for insertion into a container of a glass probe analyte sensor.
[0077] In an embodiment, the analyte is hydrogen ion and the
redox-active moiety is sensitive to hydrogen ion concentration. In
another embodiment, the sensor comprises a plurality of
redox-active moieties, wherein at least one of the redox-active
moieties is sensitive to the presence of an analyte, and at least
one other redox-active moiety is substantially insensitive to the
presence of the analyte. In another embodiment, the analyte is
hydrogen ion, and the moiety that is substantially insensitive to
the presence of hydrogen ion has a substituent selected from the
group consisting of ferrocene, polyvinylferrocene, viologen,
polyviologen, and polythiophene. In another embodiment, the moiety
that is substantially insensitive to the presence of hydrogen ion
is ferrocene or a derivative of ferrocene. In another embodiment,
the analyte is hydrogen, and the redox-active moiety that is
sensitive to the presence of the hydrogen ion comprises a
substituent selected from the group consisting of anthracenes,
quinones, anthroquinones, phenanthroquinones, phenylene diamines,
catechols, phenothiazinium dyes, monoquaternized
N-alkyl-4,4'-bipyridinium, RuOx, and Ni(OH).sub.2. In another
embodiment, the redox-active moiety that is sensitive to the
presence of hydrogen ion comprises a substituent comprising
anthracene. In another embodiment, the redox-active moiety that is
sensitive to the presence of hydrogen ion comprises a substituent
comprising an anthraquinone or a phenanthraquinone. In another
embodiment, the redox-active moiety that is sensitive to the
presence of an analyte is sensitive to the concentration of the
analyte. In another embodiment, the oxidation potential and/or
reduction potential of the redox-active moiety is sensitive to the
concentration of the analyte in a range from 10.sup.-3 M to
10.sup.-10 M. In another embodiment, the oxidation potential and/or
reduction potential of the redox-active moiety is sensitive to the
concentration of the analyte in a range from 10.sup.-1 M to
10.sup.-14 M.
[0078] In an embodiment, the sensor detects hydrogen ion
concentration from pH 1 to pH 14. In another embodiment, the sensor
detects hydrogen ion concentration from pH 3 to pH 10. In another
embodiment, the sensor measures hydrogen ion concentration within
an accuracy of plus or minus 0.1 pH units. In another embodiment,
the sensor measures hydrogen ion concentration within an accuracy
of plus or minus 0.01 pH units.
[0079] In an embodiment, the redox-active moiety is covalently
bound to the surface of the electrode, such as through an
oxygen-to-surface, carbon-to-surface or sulfur-to-surface bond. In
another embodiment, the redox-active moiety is bound to a polymer
that is immobilized onto the surface of the electrode.
[0080] In an embodiment, the semiconductor electrode is doped. In
another embodiment, the semiconductor electrode is p-doped. In
another embodiment, the semiconductor electrode is doped with
boron. In another embodiment, the semiconductor electrode is
n-doped. In another embodiment, the semiconductor electrode is
doped with phosphorous. In another embodiment, the semiconductor
electrode comprises a monolithic piece of silicon. In another
embodiment, the semiconductor electrode comprises a composite
electrode, the composite electrode comprising semiconductor
particles in a matrix. In another embodiment, the semiconductor
electrode comprises a composite electrode bound to a conductive
substrate. In another embodiment, the semiconductor particles are
present in an amount such that a conductive path is created across
the composite electrode. In another embodiment, the electrode
comprises silicon. In another embodiment, the electrode comprises
unpolished silicon. In another embodiment, the electrode comprises
polished silicon. In another embodiment, the semiconductor
electrode has a resistivity within the range of about 0.01 to 1000
.OMEGA.-cm. In another embodiment, the semiconductor electrode has
a resistivity within the range of 1 to 100 .OMEGA.-cm.
[0081] In an embodiment, the sensor is capable of measuring analyte
concentration without calibration with an external standard. In
another embodiment, the sensor remains sensitive to the analyte
without calibration after a first use by an end user. In another
embodiment, the analyte is hydrogen ion and the sensor remains
sensitive to hydrogen ion after exposure to a cell culture medium
for at least 6 days. In another embodiment, the sensor is capable
of measuring pH with an accuracy of plus or minus 0.2 pH units
after exposure to the cell culture medium for at least 6 days. In
another embodiment, the analyte is hydrogen ion and the sensor is
capable of measuring pH with an accuracy of plus or minus 0.2 pH
units after autoclave treatment at 121.degree. C. for 40 minutes.
In another embodiment, the sensor is capable of measuring pH with
an accuracy of plus or minus 0.2 pH units after autoclave treatment
at 121.degree. C. for 400 minutes. In another embodiment, the
semiconductor substrate has a plurality of zones, wherein at least
a first zone is sensitive to an analyte and a second zone is
insensitive to an analyte.
[0082] In an embodiment, the container is cylindrical in shape. In
another embodiment, the container has a circular cross-section. In
another embodiment, the container is formed of one or more metals.
In another embodiment, the one or more metals include stainless
steel.
[0083] Another aspect of the invention provides a sensor comprising
a solid state electrode having a surface immobilized thereon a
mixed layer of hydrocarbon molecules and redox-active moieties. In
an embodiment, the redox-active moieties are sensitive to an
analyte, such as hydrogen. In another embodiment, the redox-active
moieties are insensitive to the analyte.
[0084] Another aspect of the invention provides a sensor for
detecting the presence or absence of an analyte, comprising a
semiconductor electrode having a surface having immobilized thereon
a layer of a redox-active moiety, wherein the redox-active moiety
exhibits an oxidation potential and/or a reduction potential that
is sensitive or insensitive to the presence of the analyte. The
sensor further comprises a light blocking layer adjacent to the
layer of the redox-active moiety. In an embodiment, the light
blocking layer comprises a polymeric material. In another
embodiment, the polymeric material comprises a
fluoropolymer-copolymer. In another embodiment, the polymeric
material comprises Nafion. In another embodiment, the sensor
further comprises a protective layer adjacent to the light blocking
layer. In another embodiment, the protective layer comprises a
polymeric material. In another embodiment, the polymeric material
comprises polyethersulphone.
[0085] In an embodiment, the light blocking layer comprises a
composite material. In another embodiment, the composite material
comprises Nafion. In another embodiment, the composite material
comprises a porous plastic. In another embodiment, the composite
material is a porous membrane.
[0086] Another aspect of the invention provides a redox-active
moiety-containing analyte sensor for use with a glass probe analyte
detection system. The redox-active moiety-containing analyte sensor
can have any of the features and characteristics of sensors
described above.
[0087] Another aspect of the invention provides a redox-active
moiety-containing analyte sensor for use in a time period of about
1 day, or 5 days, or 10 days, or 20 days, or 25 days, or 30 days.
The redox-active moiety-containing analyte sensor can have any of
the features and characteristics of sensors described above.
[0088] Another aspect of the invention provides a redox-active
moiety-containing analyte sensor having a sensitivity between about
20 mV per pH unit and 60 mV per pH unit. The redox-active
moiety-containing analyte sensor can have any of the features and
characteristics of sensors described above.
[0089] Another aspect of the invention provides a redox-active
moiety-containing analyte sensor having a shelf life between about
3 months and 3 years. The redox-active moiety-containing analyte
sensor can have any of the features and characteristics of sensors
described above.
[0090] Another aspect of the invention provides a sensor for
detecting an analyte having an accuracy to within 0.001 pH units
while in use or storage for at least 2 years. The redox-active
moiety-containing analyte sensor can have any of the features and
characteristics of sensors described above.
[0091] Another aspect of the invention provides a sensor for
detecting an analyte having an accuracy to within 0.001 pH units
while in use or storage for at least 4 years. The redox-active
moiety-containing analyte sensor can have any of the features and
characteristics of sensors described above.
[0092] Another aspect of the invention provides a sensor for
detecting an analyte having an accuracy to within 0.001 pH units
while in use or storage for at least 8 years. The redox-active
moiety-containing analyte sensor can have any of the features and
characteristics of sensors described above.
[0093] Another aspect of the invention provides a sensor for
detecting an analyte having an accuracy to within 0.001 pH units
while in use or storage for at least 10 years. The redox-active
moiety-containing analyte sensor can have any of the features and
characteristics of sensors described above.
[0094] Another aspect of the invention provides a sensor for
detecting the presence or absence of an analyte, comprising an
electrode having a surface having immobilized thereon a
redox-active moiety, wherein the redox-active moiety exhibits an
oxidation potential and/or a reduction potential that is sensitive
or insensitive to the presence of the analyte, and a light-emitting
device adjacent to the electrode, the light-emitting device
configured to generate light. In an embodiment, the light-emitting
device is configured to generate light that is i) incident on the
surface, ii) incident on another surface of the electrode, the
another surface opposite from the surface, and/or iii) directed
through the electrode. In another embodiment, the electrode is
formed of a solid state material. In another embodiment, the
electrode is formed of a semiconductor material. In another
embodiment, the semiconductor material includes silicon. In another
embodiment, the electrode is formed of a non-semiconductor
material. In another embodiment, the non-semiconductor material
includes carbon. In another embodiment, the light-emitting device
is a light-emitting diode having an active region configured to
generate light upon the recombination of electrons and holes. In
another embodiment, the light-emitting diode is an organic
light-emitting diode. In another embodiment, during use, light from
the light-emitting device is incident on the surface.
[0095] Another aspect of the invention provides a sensor for
detecting the presence or absence of an analyte, comprising a
working electrode having a redox active moiety formed adjacent a
light emitting device. The sensor can have any of the features and
characteristics of sensors described above.
[0096] Another aspect of the invention provides a solid state
sensor for detecting the presence or absence of an analyte,
comprising a solid state electrode and a light blocking layer
adjacent to the solid state electrode. In an embodiment, the light
blocking layer is formed of a polymeric material. In another
embodiment, the light blocking layer transmits less than 10% of
light incident on the light blocking layer. In another embodiment,
the light blocking layer transmits less than 5% of light incident
on the light blocking layer. In another embodiment, the light
blocking layer transmits less than 1% of light incident on the
light blocking layer.
[0097] Another aspect of the invention provides a solid state
sensor for detecting the presence or absence of an analyte,
comprising a solid state electrode configured to detect the
presence or absence of the analyte, and a light emitting device
adjacent to the solid state electrode. The sensor can have any of
the features and characteristics of sensors described above.
[0098] Another aspect of the invention provides a sensor for
detecting the presence or absence of an analyte, comprising a
working electrode having a redox active moiety formed adjacent a
light emitting device. The sensor can have any of the features and
characteristics of sensors described above.
[0099] Another aspect of the invention provides a method for
detecting the presence or absence of an analyte, comprising
bringing an analyte sensor in contact with a sample, the analyte
sensor having an electrode having immobilized thereon a
redox-active moiety. The redox-active moiety exhibits an oxidation
potential and/or a reduction potential that is sensitive to the
presence of the analyte. Next, with the aid of the analyte sensor,
the analyte is detected at an accuracy within at least about 5%
without re-calibration for a period of at least about 1 day. In an
embodiment, the accuracy is within at least about 1%. In another
embodiment, the accuracy is within at least about 0.1%. In another
embodiment, the period is at least about 7 days. In another
embodiment, the period is at least about 1 month. In another
embodiment, the period is at least about 1 year. In another
embodiment, the period is at least about 2 years.
[0100] Another aspect of the invention provides a method for
detecting the presence or absence of an analyte, comprising using a
sensor, as described above, to detect the presence or absence of
the analyte. In some embodiments, the analyte is hydrogen ion.
[0101] Another aspect of the invention provides a sensor having a
solid state working electrode having disposed thereon a
redox-active moiety exhibiting an oxidation potential and/or a
reduction potential that is sensitive to the presence of an
analyte. The working electrode has a size and shape for use in
glass probe sensor, a reactor, a flow system, or a sample
separation system. In an embodiment, the reactor is a bioreactor.
In another embodiment, the sensor further comprises an additional
working electrode having disposed thereon a redox-active moiety
exhibiting an oxidation potential and/or reduction potential that
is insensitive to the presence of the analyte. In another
embodiment, the working electrode is doped p-type and the
additional working electrode is doped n-type or p-type. In another
embodiment, the working electrode has a resistivity is greater than
or equal to about 1 .OMEGA.-cm and the additional working electrode
has a resistivity greater than or equal to about 5
.mu..OMEGA.-cm.
[0102] Another aspect of the invention provides an analyte sensor,
comprising a first solid state working electrode and a second solid
state working electrode. The first solid state working electrode
has disposed thereon a redox-active moiety exhibiting an oxidation
potential and/or a reduction potential that is sensitive to the
presence of an analyte, the first solid state working electrode
doped p-type. The second solid state working electrode has disposed
thereon a redox-active moiety exhibiting an oxidation potential
and/or a reduction potential that is insensitive to the presence of
the analyte, the second solid state working electrode doped n-type
or p-type. In an embodiment, the first solid state working
electrode is disposed adjacent to the second solid state working
electrode. In another embodiment, the first solid state working
electrode is electrically isolated from the second solid state
working electrode. In another embodiment, the first solid state
working electrode has a resistivity greater than or equal to 1
.OMEGA.-cm (also ".OMEGA.cm" herein). In another embodiment, the
second solid state working electrode has a resistivity greater than
or equal to about 5 .mu..OMEGA.-cm. In another embodiment, the
first and second solid state working electrodes are formed of a
semiconductor. In another embodiment, the semiconductor is silicon.
In another embodiment, the second solid state working electrode is
doped n-type and has a resistivity greater than or equal to about 1
.OMEGA.-cm. In another embodiment, the resistivity of the second
solid state working electrode is between about 1 .OMEGA.-cm and 90
.OMEGA.-cm. In another embodiment, the solid state working
electrodes are disposed on a substantially flat surface of the
analyte sensor.
[0103] Another aspect of the invention provides a method for
forming an analyte sensor, comprising inserting a sensor as
described herein into a container that is for use with a glass
probe analyte sensor. In an embodiment, the method further
comprises removing a glass probe analyte sensor from the container
prior to inserting the sensor into the container.
[0104] Another aspect of the invention provides a method for
forming an analyte sensor, comprising inserting a sensor as
described herein into a container that is for use with a reactor,
in-line flow system, or sample preparation, or analysis.
[0105] Additional aspects and advantages of the present disclosure
will become readily apparent to those skilled in this art from the
following detailed description, wherein only illustrative
embodiments of the present disclosure are shown and described. As
will be realized, the present disclosure is capable of other and
different embodiments, and its several details are capable of
modifications in various obvious respects, all without departing
from the disclosure. Accordingly, the drawings and description are
to be regarded as illustrative in nature, and not as
restrictive.
INCORPORATION BY REFERENCE
[0106] All publications and patent applications mentioned in this
specification are herein incorporated by reference to the same
extent as if each individual publication or patent application was
specifically and individually indicated to be incorporated by
reference.
BRIEF DESCRIPTION OF THE DRAWINGS
[0107] The features and advantages of the invention may be further
explained by reference to the following detailed description and
accompanying drawings (or figures, also "Fig." and "FIG." herein)
that sets forth illustrative embodiments.
[0108] FIG. 1(a) shows a blow up drawing illustrating an embodiment
of the invention comprising a semiconductor electrode sensor in a
housing assembly, in accordance with an embodiment of the
invention. FIG. 1(b) shows an exemplary housing assembly comprising
the semiconductor electrode sensor, in accordance with an
embodiment of the invention;
[0109] FIG. 2 depicts an embodiment of the invention comprising a
unit that electrically connects to the semiconductor electrode
sensor and comprising a source for supplying a plurality of
potentials and a current measuring device, in accordance with an
embodiment of the invention
[0110] FIG. 3 depicts an embodiment of the invention comprising a
probe for measuring analytes within a reactor comprising two
working silicon electrodes, in accordance with an embodiment of the
invention;
[0111] FIG. 4 illustrates a method of preparing of H-terminated
silicon surface (Si--H), in accordance with an embodiment of the
invention;
[0112] FIG. 5 illustrates a silicon surface derivatization with
ferrocene moieties, vinyl-ferrocene (VFc) and ferrocene
carboxaldehyde (FcA) by covalent attachment, in accordance with an
embodiment of the invention;
[0113] FIG. 6 illustrates a silicon surface derivatized with
anthracene moieties, vinyl anthracene (VA) and anthraldehyde (AnA)
by covalent attachment, in accordance with an embodiment of the
invention;
[0114] FIG. 7 illustrates a silicon surface derivatized with both
the anthracene (VA) and ferrocene (VFc) moieties by covalent
attachment, in accordance with an embodiment of the invention;
[0115] FIG. 8 depicts a schematic diagram and picture of an
exemplary electrochemical cell, in accordance with an embodiment of
the invention;
[0116] FIG. 9 depicts square wave voltammograms showing the effect
of pH on VFc derivatized silicon sample at pH solution of 1.23,
4.61, 7.33 and 9.33, in accordance with an embodiment of the
invention;
[0117] FIG. 10(a) depicts square wave voltammograms showing the
effect of pH on VA derivatized silicon sample at pH solutions of
1.23, 4.61, 7.33 (not shown; peak maximum between -0.4 and -0.6 V,
to the left of the pH 4.61 peak and to the right of the pH 13.63
peak) and 13.63, in accordance with an embodiment of the invention.
FIG. 10(b) depicts a plot of peak potential against pH using the VA
derivatized silicon sample, in accordance with an embodiment of the
invention;
[0118] FIG. 11 depicts: (a) square wave voltammograms showing the
effect of pH on VA+VFc derivatized silicon sample at pH solutions
of 1.23, 4.65, 5.52 and 9.32 (VA circle, peaks going from right to
left); and (b) a plot of peak potential difference against pH using
the VA+VFc derivatized silicon sample, in accordance with an
embodiment of the invention;
[0119] FIG. 12 depicts square wave voltammograms showing the effect
of 10 autoclave cycles on FcA+VA derivatized silicon sample, in
accordance with an embodiment of the invention. The electrochemical
measurements were conducted at pH 6.52 buffer prior to autoclave
and after autoclave;
[0120] FIG. 13 depicts square wave voltammograms showing minimal
fouling on FcA+AnA derivatized silicon samples, in accordance with
an embodiment of the invention. The electrochemical measurements
were conducted at pH 6.52 buffer before and after six days exposure
in the cell culture at four different FcA+AnA derivatized silicon
samples (a), (b), (c) and (d);
[0121] FIG. 14 depicts a square wave voltammogram obtained at
FcA+AnA derivatized silicon sample in cell culture medium after
sterilization and 6 days exposure, in accordance with an embodiment
of the invention;
[0122] FIG. 15(a) depicts square wave voltammetric responses FcA on
Si(100, N-type, 1-5 m.OMEGA. cm) in pH 7.33 buffer medium, showing
every 50.sup.th scan of the 2,500 consecutive runs, in accordance
with an embodiment of the invention. FIG. 15(b) depicts
voltammetric responses of VFc on Si(111, N-type, 0.02-0.05
.OMEGA.cm in pH 7.33 buffer medium, showing every 50.sup.th scan of
the 2,500 consecutive runs, in accordance with an embodiment of the
invention;
[0123] FIG. 16 depicts the square wave voltammetric response of Ac
derivatized silicon surface at various temperatures (8, 17, 28 44,
56.degree. C.) in pH 7.33 buffer medium, in accordance with an
embodiment of the invention;
[0124] FIG. 17 is a drawing of an embodiment of a bioreactor of the
invention comprising a silicon-based voltammetric sensor, in
accordance with an embodiment of the invention;
[0125] FIG. 18(a) depicts voltammograms taken on an anthracene
derivatized silicon sensor over the 7 day period in cell culture
medium (every 250.sup.th scan of the 10,000 consecutive runs), in
accordance with an embodiment of the invention. FIG. 18(b) depicts
a plot of the anthracene peak potential over the 7 day time period,
in accordance with an embodiment of the invention;
[0126] FIG. 19 has charts showing the peak current of silicon
substrates derivatized with (a) vinyl ferrocene and (b) ferrocene
carboxaldehyde in pH 1.63 solution for four types of doped silicon,
in accordance with an embodiment of the invention;
[0127] FIG. 20 depicts square wave voltammograms obtained with a
four electrode system having highly-doped N-type silicon
derivatized with vinyl ferrocene and at a lightly-doped p-type
silicon derivatized with anthracene carboxaldehyde, in accordance
with an embodiment of the invention;
[0128] FIG. 21 depicts an exemplary embodiment of a probe of the
present invention having a four electrode configuration with a ring
reference electrode (RE) and a ring counter electrode (CE), in
accordance with an embodiment of the invention. FIG. 21(a) is a
shaded drawing, and FIG. 21(b) is a line drawing;
[0129] FIG. 22A schematically illustrates an electrochemical sensor
having three modules; FIG. 22B is an enlarged view of a portion of
the electrochemical sensor of FIG. 22A, in accordance with an
embodiment of the invention;
[0130] FIG. 23 schematically illustrates a printed circuit board,
in accordance with an embodiment of the invention;
[0131] FIG. 24 schematically illustrates a sensor formed on a
printed circuit board and mounted on a head assembly, in accordance
with an embodiment of the invention;
[0132] FIG. 25A schematically illustrates an electrochemical sensor
mounted on a wall of a disposable container, in accordance with an
embodiment of the invention; FIG. 25B illustrates an
electrochemical sensor comprising electrodes formed on a printed
circuit board, in accordance with an embodiment of the
invention;
[0133] FIG. 26 schematically illustrates an electrochemical sensor
mounted on a chamber of a flow-through tube, in accordance with an
embodiment of the invention;
[0134] FIG. 27A schematically illustrates an electronics unit for
permitting integration of an electrochemical sensor into current
probe systems, in accordance with an embodiment of the invention;
FIG. 27B illustrates a probe attached to the electronics unit of
FIG. 27A, in accordance with an embodiment of the invention; FIG.
27C shows a probe attached to an electronic unit, which is in turn
attached to a reader, in accordance with an embodiment of the
invention;
[0135] FIG. 28 schematically illustrates an electrochemical probe
mounted on a bioreactor, in accordance with an embodiment of the
invention;
[0136] FIG. 29 shows a working electrode having a light-emitting
device and an electrode adjacent to the light-emitting device, in
accordance with an embodiment of the invention;
[0137] FIG. 30 shows sensor output (y-axis, mV) with time (x-axis)
during pH measurements for a shielded (top) and unshielded (bottom)
sensors;
[0138] FIGS. 31A-31F show sensor output (y-axis; current, arbitrary
units) at various pH's and under light and dark conditions as a
function of voltage (mV);
[0139] FIGS. 32A-32E show exemplary sensors having form factors
suited for various applications; and
[0140] FIG. 33 shows the pH of a fermentation reactor as a function
of time as measured by a glass electrode sensor and a redox-active
moiety-based pH sensor.
DETAILED DESCRIPTION OF THE INVENTION
[0141] While preferable embodiments of the invention have been
shown and described herein, it will be obvious to those skilled in
the art that such embodiments are provided by way of example only.
Numerous variations, changes, and substitutions will now occur to
those skilled in the art without departing from the invention. It
should be understood that various alternatives to the embodiments
of the invention described herein may be employed in practicing the
invention.
[0142] The invention relates to compositions, devices, systems, and
methods for producing and using solid state electrodes modified
with redox-active agents as sensors. The subject devices and
systems are particularly useful for voltammetrically measuring
concentrations of an analyte of interest. The sensors of the
present invention utilize a solid state electrode comprising
redox-active species on its surface. At least one redox-active
species on the solid sate (e.g., semiconductor) surface has a redox
potential (reduction potential or oxidation potential) that is
sensitive to the presence and or amount of an analyte of interest.
Voltammetry can be performed on the solid state electrode and used
to measure the redox potential of the analyte-sensitive redox
groups on the surface of the electrode. The measured value of the
redox potential can then be used to determine the concentration of
an analyte, for example an analyte in solution. In some
embodiments, the solid state electrode of the present invention has
more than one redox-active species. In an embodiment, at least one
redox-active species is sensitive to the presence of an analyte
(e.g., protons) and another redox-active species is insensitive to
the presence of the analyte. Another aspect of the invention
relates to the measurement of the concentration of hydrogen ion, or
pH using the subject devices or systems. The surface modified solid
state sensors of the present invention can be used to measure the
pH of a variety of solutions. The surface modified sensors of the
present invention are robust, reliable, accurate, and/or can be
made such that they do not require calibration.
[0143] In some embodiments, solid state electrodes are formed of
semiconductors. Semiconductors have advantages as substrates and
sensors for the methods of the present invention. Semiconductors
have band gaps that may be modified with the aid of chemical
dopants, which can aid in preparing sensitive and accurate sensors.
In addition, semiconductors can be less prone to fouling and
degradation than other substrates.
[0144] Semiconductor surfaces, for example, inorganic
semiconductors such as silicon and organic semiconductors, can be
amenable to surface modification, e.g., covalent modification.
Semiconductors generally have electronic band structures, the
characteristics of which can be modified, for example, by doping.
In some cases, the semiconductor that is used is silicon. An
advantage of using silicon as a substrate and as an electrode is
that silicon is amenable to mass production. In particular,
semiconductor processing techniques are readily available for
producing silicon electrodes in large quantities at low cost. In
addition, existing semiconductor processing techniques make it
feasible to integrate electronic functionality into the material
comprising the silicon electrode. Another advantage of silicon is
that it can form strong covalent bonds, for example with carbon,
nitrogen, oxygen, thus allowing for the facile and robust
modification of the surface in a manner required for its intended
uses. For instance, a silicon surface can be modified to attachment
of any suitable redox-active agent. Silicon is also an advantageous
surface for carrying out voltammetry in that it is stable to a wide
range of electrical potential without undergoing degradation.
[0145] An aspect of the invention is a surface modified solid state
(e.g., semiconductor) redox sensor that can measure analyte
concentration reliably and consistently with minimal intervention,
such as re-calibration. Another aspect of the invention is a
semiconductor redox sensor that does not require calibration or
re-calibration. The ability to sense analytes, such as hydrogen ion
without calibration has a number of advantages for analyte
measurements, for example, for in-line monitoring. For example, it
allows for ease of operator handling for single point measurements.
In some embodiments, the sensors of the present invention are
included in in-line operator-independent control measurements. Such
in-line measurements can be made independent of the operator and
can be used for process control, for example for pH measurements
for process control. The subject sensors can be set to provide
real-time measurements of an analyte, including, but not limited to
real time-measurements of hydrogen ion concentration.
Semiconductor Substrates
[0146] An aspect of the invention provides an electrochemical
sensor having a working electrode formed of a semiconductor
substrate. The semiconductor substrate can comprise any suitable
semiconductor material including those known in the art and those
described herein. The semiconductor substrate can be an inorganic
semiconductor or an organic semiconductor. The semiconductor
substrate can be doped or undoped. In some embodiments the
semiconductor substrate comprises silicon.
[0147] A semiconductor substrate of the invention is generally a
solid material that has electrical conductivity in between that of
a conductor and that of an insulator. The conductivity can vary
over that wide range either permanently or dynamically.
[0148] Inorganic semiconductor substrates of the invention can
comprise, for example, Group IV elemental semiconductors, such as
diamond (C), silicon (Si), germanium (Ge); Group IV compound
semiconductors, such as silicon carbide (SiC), silicon nitride
(SiN), silicon germanide (SiGe), Group III-V semiconductors, such
as aluminum antimonide (AlSb), aluminum arsenide (AlAs), aluminum
nitride (AlN), aluminum phosphide (AlP), boron nitride (BN), boron
phosphide (BP), boron arsenide (BAs), gallium antimonide (GaSb),
gallium arsenide (GaAs), gallium nitride (GaN), gallium phosphide
(GaP), indium antimonide (InSb), indium arsenide (InAs), indium
nitride (InN), indium phosphide (InP); Group III-V ternary
semiconductor alloys, such as aluminum gallium arsenide (AlGaAs,
AlxGal-xAs), indium gallium arsenide (InGaAs, InxGal-xAs), indium
gallium phosphide (InGaP), aluminum indium arsenide (AlInAs),
aluminum indium antimonide (AlInSb), gallium arsenide nitride
(GaAsN), gallium arsenide phosphide (GaAsP), aluminum gallium
nitride (AlGaN), aluminum gallium phosphide (AlGaP), indium gallium
nitride (InGaN), indium arsenide antimonide (InAsSb), indium
gallium antimonide (InGaSb), Group III-V quaternary semiconductor
alloys, such as aluminum gallium indium phosphide (AlGaInP, also
InAlGaP, InGaAlP, AlInGaP), aluminum gallium arsenide phosphide
(AlGaAsP), indium gallium arsenide phosphide (InGaAsP), Aluminum
indium arsenide phosphide (AlInAsP), aluminum gallium arsenide
nitride (AlGaAsN), indium gallium arsenide nitride (InGaAsN),
indium aluminum arsenide nitride (InAlAsN), gallium arsenide
antimonide nitride (GaAsSbN), Group III-V quinary semiconductor
alloys, such as gallium indium nitride arsenide antimonide
(GaInNAsSb), gallium indium arsenide antimonide phosphide
(GaInAsSbP), II-VI semiconductors, cadmium selenide (CdSe), cadmium
sulfide (CdS), cadmium telluride (CdTe), zinc oxide (ZnO), zinc
selenide (ZnSe), zinc sulfide (ZnS), zinc telluride (ZnTe), Group
II-VI ternary alloy semiconductors, such as cadmium zinc telluride
(CdZnTe, CZT), mercury cadmium telluride (HgCdTe), mercury zinc
telluride (HgZnTe), mercury zinc selenide (HgZnSe), Group I-VII
semiconductors, such as cuprous chloride (CuCl), Group IV-VI
semiconductors, such as lead selenide (PbSe), lead sulfide (PbS),
lead telluride (PbTe), tin sulfide (SnS), tin telluride (SnTe),
Group IV-VI ternary semiconductors, such as lead tin telluride
(PbSnTe), Thallium tin telluride (Tl2SnTe5), thallium germanium
telluride (Tl2GeTe5), Group V-VI semiconductors, such as bismuth
telluride (Bi2Te3), and Group II-V semiconductors, such as cadmium
phosphide (Cd3P2), cadmium arsenide (Cd3As2), cadmium antimonide
(Cd3Sb2), zinc phosphide (Zn3P2), zinc arsenide (Zn3As2), and zinc
antimonide (Zn3 Sb2).
[0149] The inorganic semiconductor substrates of the invention can
also comprise layered semiconductors, such as lead(II) iodide
(PbI2), molybdenum disulfide (MoS2), gallium selenide (GaSe), tin
sulfide (SnS), bismuth sulfide (Bi2S3), other semiconductors, such
as copper indium gallium selenide (CIGS), platinum silicide (PtSi),
bismuth(III) iodide (BiI3), mercury(II) iodide (HgI2), thallium(I)
bromide (TlBr), and miscellaneous oxides, such as titanium dioxide:
anatase (TiO2), copper(I) oxide (CuO0), copper(II) oxide (CuO),
uranium dioxide (UO2), and uranium trioxide (UO3).
[0150] In some embodiments of the invention, the semiconductor
substrate can comprise an organic semiconductor. The organic
semiconductor is any suitable organic material that has
semiconductor properties. The organic semiconductor substrates of
the invention can comprise, for example, small molecules, short
chain (oligomers) and long chain (polymers). Examples of
semiconducting small molecules (e.g., unsaturated and aromatic
hydrocarbons) are pentacene, anthracene, rubrene, tetracene,
chrysene, pyrene, perylene, coronene; metal complexes of porphine
and phthalocyanine, compounds such as zinc
1,10,15,20-tetraphenyl-21H,23H-porphine, copper phthalocyanine,
lutetium bisphthalocyanine, and aluminum phthalocyanine chloride
can be used. Suitable derivatives of these small molecules can also
be used. In some embodiments, the organic semiconductor substrates
of the invention can comprise thin films.
[0151] Examples of semiconducting polymers or oligomers include
suitable conjugated hydrocarbon or heterocyclic polymers or
oligomers. Suitable polymers or oligomers include: polyaniline,
polypyrrole, and polythiophene, poly(3-hexylthiophene),
poly(p-phenylene vinylene), F8BT, polyacetylene, polydiacetylene,
polyacene, polyphenylene, poly(phenylene vinylene), polyfuran,
polypyridine, poly(thienylene vinylene), poly(ferrocenyl vinylene
phenylene vinylene), poly(fluorine), and poly(carbazole), and
combinations thereof. Derivatives of these polymers, for instance
derivatives having functional side chains amenable to the
attachment of redox active species, can be used.
[0152] Other examples of semiconducting polymers are
poly(anilinesulfonic acid), poly(ferrocenyl vinylene phenylene
vinylene), poly(fluorine), and poly(carbazole). The organic
semiconductor substrates of the invention can comprise, for
example, organic charge-transfer complexes, and "linear backbone"
polymers derived from polyacetylene, such as polyacetylene
polypyrrole, and polyaniline. Charge-transfer complexes can exhibit
similar conduction mechanisms to inorganic semiconductors. This
includes the presence of a hole and electron conduction layer and a
band gap. The materials can exhibit tunneling, localized states,
mobility gaps, and phonon-assisted hopping. Organic semiconductors
can be doped. In some embodiments, the invention can utilize highly
doped organic semiconductors, for example Polyaniline (Ormecon) and
PEDOT:PSS. In some cases organic semiconductors can be produced
such that they are transparent and/or flexible, which can be useful
in some embodiments.
[0153] The semiconductors of the invention can typically be
characterized as having a band gap, the band gap representing the
amount of energy separating the valence and conduction bands of the
semiconductor. The addition of dopants makes the band gap smaller,
tending to allow more facile promotion of electrons from the
valence band to the conduction band. A smaller band gap can result
in higher conductivity for the semiconductor substrate. The band
gap and conductivity characteristics of the semiconductor
substrates can be controlled in some cases by the introduction of
dopants. In some cases upon the addition of a sufficiently large
proportion of dopants, the semiconductor substrates of the
invention can conduct electricity nearly as well as metals.
Depending on the kind of dopant or impurity, a doped region of
semiconductor can have more electrons or holes, and is named N-type
or P-type (herein also "n-type" and "p-type") semiconductor
material, respectively. Junctions between regions of N- and P-type
semiconductors create electric fields, which cause electrons and
holes to be available to move away from them, and this effect is
critical to semiconductor device operation. Also, a density
difference in the amount of impurities produces a small electric
field in the region which is used to accelerate non-equilibrium
electrons or holes.
[0154] In some embodiments the presence of the band gap can be
advantageous. For example, when performing electrochemistry, when
the Fermi energy of a doped semiconductor lies at the same energy
as the solution/molecular redox potential at a certain potential,
generally no net transfer of charge/current will flow from the
redox species (immobilized on the surface or in the solution) to
the substrate or from the substrate to the redox species. This
potential is sometimes referred to as the flatband potential. The
location of the flatband potential can be influenced by dopant
densities. The Mott-Schottky equation (see P. Schmuki, H. Bohni and
J. A. Bardwell, J. Electrochem. Soc., 1995, 142, 1705) can be used
to estimate the flat band potential.
[0155] A conductor electrode generally does not have a flatband
potential thus has a broad potential window where current can flow
from the redox species to the substrate. For example, when the
conductor electrode is exposed to an aqueous solution containing a
mixture of several redox active species, currents corresponding to
these redox species (i.e., non-specific adsorption) may be recorded
unless additional efforts are made to screen off these non-specific
interactions, e.g., putting a diluent layer onto the conductor
electrode. The non-specific interactions can be reduced or
eliminated by using a semiconductor electrode wherein the
semiconductor has a the band gap that has a limited potential
window, such that the semiconductor can only conduct current for
the electrochemical reaction occurring within that limited window.
For example, the limited window can be between -1.0 to 0 V. Thus,
in some embodiments, non-specific interactions in the solution can
be reduced or eliminated by using a semiconductor electrode of the
invention with the appropriate dopant density (which can be
estimated using the Mott-Schottky equation).
[0156] One useful aspect of the semiconductors of the invention is
that their conductivity can be modified by introducing impurities
(dopants) into their crystalline lattice or amorphous regions. The
process of adding controlled impurities to a semiconductor can be
referred to as doping. The amount of impurity, or dopant, added to
an intrinsic (pure) semiconductor alters its level of conductivity.
Doped semiconductors may be referred to as extrinsic. The
semiconductors of the present invention can be either intrinsic or
extrinsic semiconductors.
[0157] Suitable dopants can be chosen, as is known in the art, on
the atomic properties of the dopant and the material to be doped.
In general, dopants that produce the desired controlled changes are
classified as either electron acceptors or donors. A donor atom
that activates (e.g., becomes incorporated into the crystal
lattice) donates weakly-bound valence electrons to the material,
creating excess negative charge carriers. These weakly-bound
electrons can move about in the semiconductor relatively freely and
thus can facilitate electrical conduction in the presence of an
electric field. In some cases, the donor atoms introduce some
states below, but very close to, the conduction band edge.
Electrons at these states can be thermally excited to conduction
band, becoming free electrons, in some cases, at room temperature.
In some embodiments an activated acceptor dopant is utilized. The
activated acceptor can produce a hole. Semiconductors doped with
donor impurities are typically called N-type, while those doped
with acceptor impurities are typically known as P-type. In some
embodiments of the invention, the semiconductor of the invention
can have both donor and acceptor dopants. In some embodiments the
semiconductor can have both n type and p type charge carriers. The
n or p type designation generally indicates which charge carrier
acts as the material's majority carrier. The opposite carrier is
generally called the minority carrier, which, in some cases, exists
due to thermal excitation at a lower concentration than the
majority carrier.
[0158] As described herein, where the semiconductor is a Group IV
semiconductor such as silicon or germanium, suitable electron
donors include, for example, phosphorous, arsenic, antimony, and
bismuth. In some embodiments the dopant is phosphorous. In some
embodiments the dopant is antimony. Where the semiconductor is a
Group IV semiconductor such as silicon or germanium, suitable
electron acceptors include boron, aluminum, gallium, and the like.
In some embodiments, the dopant is boron. Where the semiconductor
comprises an element in another group than Group IV, as is known in
the art, elements that are outside of that group can act as either
n type of p type dopants.
[0159] In addition to modification through doping, the resistance
of semiconductors can in some cases, for example, be modified
dynamically by applying electric fields. The ability to control
resistance/conductivity in semiconductor substrate or within
regions of semiconductor substrate dynamically through the
application of electric fields can be useful in some
embodiments.
[0160] The semiconductor substrate can be in any form that is
amenable to the production of a semiconductor electrode. The
semiconductor substrate can comprise a monolithic piece of the
semiconductor, a coating of the semiconductor deposited onto
another material, or a powder of semiconductor particles. The
semiconductor substrate can be a monolithic form such as a chip,
wafer, rod, needle, block, ingot, or the like. The semiconductor
substrate can alternately be in particulate form, for example in
the form of powder comprised of particles. The particles can be of
arbitrary shape or can be in the form of fibers, sheets, beads,
discs, or balls. Where the substrate is in the form of a powder
made up of particles, it will generally be formed into a composite
electrode as described in more detail below.
[0161] The semiconductor substrate of the present invention can be
a thin layer of semiconductor that is formed upon another material,
for example a thin layer of semiconductor formed on glass would
constitute a semiconductor substrate.
[0162] In an embodiment, the semiconductor substrate can include a
layer of semiconductor material having a thickness of about 0.1
nanometers ("nm") and 5000 nm, or between about 1 nm and 1000 nm,
or between about 10 nm and 500 nm. In another embodiment, the
semiconductor substrate can include a layer of silicon having a
thickness of about 0.1 nanometers ("nm") and 5000 nm, or between
about 1 nm and 1000 nm, or between about 10 nm and 500 nm.
[0163] In some embodiments the semiconductor substrate used to make
the electrode is a composite material comprising semiconductor
particles dispersed in a matrix or binder. The semiconductor
substrate can be made of a composite material comprising a powder
of semiconductor dispersed in a binder to make a composite
semiconductor substrate. The semiconductor powder can be in the
form of spheres, crystallites, rods, fibers, or any other arbitrary
shape. In an embodiment the composite electrode is made of
semiconductor crystallites dispersed in a polymeric matrix. The
matrix or binder can be an organic, inorganic, or organometallic
polymer. Non-limiting examples of useful inorganic polymeric
materials include polyphosphazenes, polysilanes, polysiloxane,
polygermanes, polymeric sulfur, polymeric selenium, silicones, and
mixtures of any of the foregoing.
[0164] In some embodiments the polymer can be an organic polymer.
Non-limiting examples of suitable organic polymeric materials
include, but are not limited to, thermoset materials and
thermoplastic materials. Non-limiting examples of polymers useful
in the invention include polyesters such as polyethylene
terephthalate, polybutylene terephthalate, and polyethylene
naphthalate, polycarbonates, polyolefins such as polyethylene,
polypropylene, and polyisobutene, acrylic polymers such as
copolymers of styrene and an acrylic acid monomer, and polymers
containing methacrylate, polyamides, thermoplastic polyurethanes,
vinyl polymers, polyimides, polyamides, polytetrafluoroethelene and
other fluoropolymers, and mixtures of any of the foregoing.
[0165] The binder can be insulating, semiconductive, or conductive.
In an embodiment, the binder is a material, such as a polymer,
which is an insulating material. Where an insulating binder is
used, the current will tend to only flow through the dispersed
semiconductor powder. In some embodiments, the binder includes
conductive components. In some embodiments, the binder comprises a
conductive polymer such as polyaniline, polyacetylene,
poly(alkylthiophene), poly(alkylpyrrole), and the like. In some
embodiments, the conductive component can comprise conductive
particles such as metal particles, such as nickel particles other
conductive particles including carbon particles. In some
embodiments, the conductive component is chosen such that the
conductive component such as the conductive polymer exhibits
reduction and/or oxidation potentials that are outside of the
reduction and/or oxidation potentials of the redox active
moieties.
[0166] The composite semiconductor substrate can be formed by
mixing the semiconductor powder with a monomer, oligomer, or
prepolymer and curing the monomer, oligomer or prepolymer to form a
polymeric matrix. The polymerization can be initiated in any manner
known in the art or disclosed herein. The polymerization can be
initiated, for example, thermally or photochemically in the
presence of an initiator. The polymerization can be carried out
with one or more crosslinkers. The cross-linkers can be chosen to
adjust the physical properties of the polymeric matrix and thus
adjust the properties of the composite semiconductor substrate. The
composite semiconductor substrate can be formed by mixing the
semiconductor powder with a molten thermoplastic polymer, forming
the substrate, and allowing the mixture to cool. The composite
semiconductor substrate can be formed by mixing the semiconductor
powder with a polymer or prepolymer in a solvent, and allowing the
solvent to evaporate to form the composite. Combinations of any of
the above methods can be used.
[0167] The electrical properties of the composite semiconductor
substrate can be affected by the amount of semiconductor, the
particle size, and the particle shape. In general, the amount of
semiconductor in the composite is high enough to create conductive
pathways throughout the material. This amount of material necessary
to provide conductive paths across the material is sometimes called
the percolation threshold. The amount of semiconductor particles
for conductivity can also depend on the processing conditions such
as the viscosity of the binder and the amount of mixing. The amount
of semiconductor is generally set at a level at which the physical
properties of the material, such as mechanical strength and
flexibility will not suffer to the point that the material is not
useful. The amount of semiconductor will generally be from 0.1
volume percent to 70 volume percent of the composite material. In
some embodiments the amount of semiconductor may be from 1 volume
percent to 50 volume percent. In some embodiments the amount of
semiconductor may be from 10 volume percent to 40 volume percent.
The amount of semiconductor can be from 1% to 5%, 5% to 10%, 10% to
15%, 10% to 20%, 20% to 30%, 30% to 40%, 40% to 50% or 50% to
60%.
[0168] The composite semiconductor substrate can be formed by
methods used for shaping polymeric materials such as coating,
molding, and casting into shapes that are useful as electrodes. The
composite semiconductor substrate electrode will generally be
connected to an electrically conductive wire to apply current and
potential. The material can be cast, coated, and/or molded onto a
conductive substrate such as a metal to form a conductive junction
for connecting conductors for transfer of current to and from the
composite electrode.
[0169] The semiconductor substrates of the invention generally have
a high enough electrically conductivity to act as electrodes, to
transmit current for the oxidation and/or reduction of the bound
redox-active moieties. To make the semiconductor substrate more
conductive, the semiconductor substrate can include impurities or
dopants to increase electrical conductivity and reduce the
resistivity.
[0170] The electrical resistivity (reciprocal of conductivity) can
be for example 0.1 (ohm-centimeters), 1 (ohm-centimeters), 10
(ohm-centimeters), 100 (ohm-centimeters), to in excess of 1000 or
even 10,000 (ohm-centimeters) or even higher which is comparable to
graphite and conventional metallic conductors.
[0171] In some embodiments the resistivity of the semiconductor
substrate is the range of 0.0001 to 100,000 .OMEGA.-cm (or
ohm-centimeters). In some embodiments the resistivity of the
semiconductor substrate is the range of 0.001 to 10,000 .OMEGA.-cm.
In some embodiments the resistivity of the semiconductor substrate
is the range of 0.01 to 1000 .OMEGA.-cm. In some embodiments the
resistivity of the semiconductor substrate is the range of 0.1 to
100 .OMEGA.-cm. In some embodiments the resistivity of the
semiconductor substrate is within the range of 1 to 100 .OMEGA.-cm.
In some embodiments the resistivity of the semiconductor substrate
is within the range of 10 to 90 .OMEGA.-cm. In some embodiments the
semiconductor substrate is single crystal semiconductor is Si(100)
that is P-type with a resistivity of 10 to 90 .OMEGA.-cm. In some
cases several semiconductor substrates with different resistivities
may be used. For example, a system of the invention may comprise
one lightly doped semiconductor substrate having one redox active
species bound to it, and also a more highly doped semiconductor
surface having another redox active species bound to it. For
example, a system of the invention may comprise one lightly doped
semiconductor surface having a pH sensitive redox active moiety
such as anthraquinone bound thereto, and a second semiconductor
surface that is more highly doped having a hydrogen ion insensitive
redox active moiety such as ferrocene bound to it.
[0172] In some cases different regions of the semiconductor
substrate can be doped at different levels. For example, it is well
known in the semiconductor processing art that a mask can be used
to cover some regions of the semiconductor, while leaving other
regions exposed. The exposed regions can be treated selectively
with dopants resulting in a semiconductor surface wherein some
regions are doped differently than other regions. By using multiple
steps with various masks, the conductivity properties of different
regions of the semiconductor surface can be controlled. Thus, some
regions can have high conductivity, some low, some regions can have
a large band gap, and other regions can have small band gaps, some
regions can have N-doping, some P-doping, and some no doping. In
addition, the various regions can be connected with conducting
regions, for example deposited metal, e.g., gold in to be able to
electrically address the various regions.
Silicon Substrates
[0173] Another aspect of the invention provides an electrochemical
sensor having a semiconductor substrate that includes silicon. The
silicon substrate can have a surface onto which are attached
redox-active moieties. The silicon substrate can comprise amorphous
silicon or silicon comprising a variety of crystalline forms. The
silicon substrate can also be polycrystalline. In some embodiments
the silicon substrate can have both amorphous and crystalline
regions. In some embodiments, the silicon can be nanocrystalline or
microcrystalline silicon. Nanocrystalline silicon and
microcrystalline silicon can be used to describe an allotropic form
of silicon with paracrystalline structure having small grains of
crystalline silicon within the amorphous phase. Where the silicon
substrate is crystalline, the surface of the silicon substrate can
have various crystalline faces on the surface. Crystalline silicon
is generally in a face centered cubic (fcc) form. In some
embodiments, such as where a polycrystalline silicon is used, the
surface may have multiple crystalline planes exposed. Where single
crystal silicon is used, in some cases, the silicon substrate can
be made to have one or more crystal planes represented or
predominantly represented on the surface. In some embodiments, the
surface of the silicon substrate will comprise one or more crystal
planes having a crystalline lattice of (xxx) wherein x is an
integer corresponding to the lattice defining the crystal plane. In
some embodiments the crystal planes (100), (010), (001), (110),
(101), or (112) may be predominantly represented at the surface. In
some embodiments a silicon substrate has the (100) plane
predominantly represented at the surface.
[0174] The silicon electrode of the present invention can comprise
a polished or an unpolished silicon substrate. Silicon is generally
polished prior to silicon processing, for example, building
features such as transistors. In some embodiments, such as those
embodiments where electronic functionality is incorporated into a
silicon sensor, a polished silicon surface may be desirable. In
other embodiments, an unpolished silicon substrate can be used. An
unpolished silicon substrate can be less expensive than a silicon
substrate that has gone through a polishing step. In addition, an
unpolished silicon substrate can have a higher surface area for a
given area of silicon than a polished silicon substrate.
[0175] The silicon electrode of the present invention can comprise
porous silicon. An advantage of porous silicon is an increase of
the effective surface area. An increased surface area can be
advantageous for providing a higher signal from the oxidation and
reduction of the surface bound redox moieties due to a higher
number of such moieties in contact with the sample. As is known in
the art, if the surface is too porous, it can become less robust.
Therefore the level of porosity can be controlled to maximize
important properties for the particular applications. The porous
silicon can be prepared by, for example, galvanostatic, chemical,
or photochemical etches from silicon wafers. In some embodiments,
chemical etching with hydrofluoric acid (HF) can be used to produce
a porous silicon substrate. In some embodiments, the average pore
size of the silicon substrate ranges from 1 nm to 500 nm. Pore size
can be measured by, for example, nitrogen gas adsorption or Hg
porosimetry. In some embodiments, the amount of porosity ranges
between 1% and 98%. In some embodiments, the amount of porosity
ranges between 5% and 75%. In some embodiments, the amount of
porosity ranges between 10% and 50%. In some embodiments, the
amount of porosity ranges between 20% and 40%. In some embodiments
the porosity is between 1% to 5%, 5% to 10%, 10% to 30%, 20% to
40%, 30% to 50%, or 40% to 60%. The porosity measurement can be
made on an area percent basis or a volume percent basis.
[0176] In addition, porous silicon could be readily integrated with
existing silicon-based integrated circuit (IC) manufacturing
processes.
[0177] The silicon substrate can be in any form that is amenable to
the production of a silicon electrode. The silicon substrate can
comprise a monolithic piece of silicon, a coating of silicon
deposited onto another material, or a powder of silicon particles.
The silicon substrate can be a monolithic form such as a chip,
wafer, rod, needle, block, ingot, or the like. The silicon
substrate can alternately be in particulate form, for example in
the form of powder comprised of particles. The particles can be of
arbitrary shape or can be in the form of fibers, sheets, beads,
discs, polyhedra, or balls. Where the substrate is in the form of a
powder made up of particles, it will generally be formed into a
composite electrode as described in more detail below.
[0178] In some embodiments the semiconductor electrode is made from
single crystal silicon. The single crystal silicon can be made by
zone melting, also called zone refining, a process in which rods of
metallurgical grade silicon are first heated to melt at one end.
Then, the heater is typically slowly moved down the length of the
rod, keeping a small length of the rod molten as the silicon cools
and re-solidifies behind it. Since most impurities tend to remain
in the molten region rather than re-solidify, when the process is
complete, most of the impurities in the rod will typically have
been moved into the end that was the last to be melted. This end is
then cut off and discarded, and the process repeated if a still
higher purity is desired. The single crystal silicon of the
invention can also be produced via the Czochralski process, (CZ-Si)
which tends to be inexpensive and is capable of producing large
size crystals.
[0179] In some embodiments the silicon electrode is
polycrystalline. As used herein, the term "polysilicon" is used
interchangeably with the term "polycrystalline silicon". In some
embodiments, the polysilicon is deposited. The polycrystalline
silicon can be deposited by low pressure chemical vapor deposition
(LPCVD), plasma-enhanced chemical vapor deposition (PECVD), or
solid-phase crystallization (SPC) of amorphous silicon in certain
processing regimes These processes can require relatively high
temperatures, for example, above 300.degree. C. The polycrystalline
silicon electrodes can also be made, for example on polymeric
substrates, using laser crystallization to crystallize a precursor
amorphous silicon (a-Si) material on a plastic substrate without
melting or damaging the plastic. In some cases, the for example,
short, high-intensity ultraviolet laser pulses are used to heat the
deposited a-Si material to above the melting point of silicon,
without melting the entire substrate. By controlling the
temperature gradients, the crystal size on the electrodes can be
controlled. Grain sizes can be, for instance from 10 nanometer to 1
micrometer. Another method to produce polysilicon at low
temperatures for the electrodes of the present invention is a
metal-induced crystallization in which an amorphous silicon thin
film is crystallized, for example at temperatures at or above
150.degree. C., while in contact of a metal film such as aluminum,
gold, or silver. The polycrystalline silicon electrodes can also be
formed onto a metal structure such as a wire. For example, the end
of a cylindrical wire can be coated with polysilicon, which can be
derivatized with redox active species as described herein. The
structure can be used as an electrode or portion of an electrode
with silicon portion accessible to the medium containing the
analyte, and the wire acting to connect the silicon electrode to
the parts of the system providing voltage and allowing for the flow
of current.
[0180] An advantage of polysilicon over amorphous silicon (a-Si) is
that the mobility of the charge carriers can be orders of magnitude
larger than in single crystal silicon and the material also can
show greater stability under electric field and light-induced
stress.
[0181] The silicon substrate of the present invention can be a thin
layer of silicon that is formed upon another material, for example
a thin layer of silicon formed on glass would constitute a silicon
substrate.
[0182] In some embodiments the silicon substrate used to make the
electrode is a composite material comprising silicon particles
dispersed in a matrix or binder. The silicon substrate can be made
of a composite material comprising a powder of silicon dispersed in
a binder to make a composite silicon substrate. The silicon powder
can be in the form of spheres, crystallites, rods, fibers, or any
other arbitrary shape. In an embodiment the composite electrode is
made of silicon crystallites dispersed in a polymeric matrix. The
matrix or binder can be an organic, inorganic, or organometallic
polymer. Non-limiting examples of useful inorganic polymeric
materials include polyphosphazenes, polysilanes, polysiloxane,
polygermanes, polymeric sulfur, polymeric selenium, silicones, and
mixtures of any of the foregoing.
[0183] In some embodiments the polymer can be an organic polymer.
Non-limiting examples of suitable organic polymeric materials
include, but are not limited to, thermoset materials and
thermoplastic materials. Non-limiting examples of polymers useful
in the invention include polyesters such as polyethylene
terephthalate, polybutylene terephthalate, and polyethylene
naphthalate, polycarbonates, polyolefins such as polyethylene,
polypropylene, and polyisobutene, acrylic polymers such as
copolymers of styrene and an acrylic acid monomer, and polymers
containing methacrylate, polyamides, thermoplastic polyurethanes,
vinyl polymers, polyimides, polyamides, polytetrafluoroethelene and
other fluoropolymers, and mixtures of any of the foregoing.
[0184] The binder can be insulating, semiconductive, or conductive.
In an embodiment, the binder is a material, such as a polymer, that
is an insulating material. Where an insulating binder is used, the
current will tend to only flow through the dispersed silicon
powder. In some embodiments, the binder includes conductive
components. In some embodiments, the binder comprises a conductive
polymer such as polyaniline, polyacetylene, poly(alkylthiophene),
poly(alkylpyrrole), and the like. In some embodiments, the
conductive component can comprise conductive particles such as
metal particles, such as nickel particles other conductive
particles including carbon particles. In some embodiments, the
conductive component is chosen such that the conductive component
such as the conductive polymer exhibits reduction and/or oxidation
potentials that are outside of the reduction and/or oxidation
potentials of the redox active moieties.
[0185] The composite silicon substrate can be formed by mixing the
silicon powder with a monomer, oligomer, or prepolymer and curing
the monomer, oligomer or prepolymer to form a polymeric matrix. The
polymerization can be initiated in any manner known in the art or
disclosed herein. The polymerization can be initiated, for example,
thermally or photochemically in the presence of an initiator. The
polymerization can be carried out with one or more crosslinkers.
The cross-linkers can be chosen to adjust the physical properties
of the polymeric matrix and thus adjust the properties of the
composite silicon substrate. The composite silicon substrate can be
formed by mixing the silicon powder with a molten thermoplastic
polymer, forming the substrate, and allowing the mixture to cool.
The composite silicon substrate can be formed by mixing the silicon
powder with a polymer or prepolymer in a solvent, and allowing the
solvent to evaporate to form the composite. Combinations of any of
the above methods can be used.
[0186] The electrical properties of the composite silicon substrate
can be affected by the amount of silicon, the particle size, and
the particle shape. In general, the amount of silicon in the
composite is high enough to create conductive pathways throughout
the material. This amount of material necessary to provide
conductive paths across the material is sometimes called the
percolation threshold. The amount of silicon particles required for
conduction can also depend on the processing conditions such as the
viscosity of the binder and the amount of mixing. The amount of
silicon is generally set at level at which the physical properties
of the material, such as mechanical strength and flexibility will
not suffer to the point that the material is not useful. The amount
of silicon will generally be from 0.1 volume percent to 70 volume
percent of the composite material. In some embodiments the amount
of silicon may be from 1 volume percent to 50 volume percent. In
some embodiments the amount of silicon may be from 10 volume
percent to 40 volume percent. The amount of silicon can be from 1%
to 5%, 5% to 10%, 10% to 15%, 10% to 20%, 20% to 30%, 30% to 40%,
40% to 50% or 50% to 60%.
[0187] The composite silicon substrate can be formed by methods
used for shaping polymeric materials such as coating, molding, and
casting into shapes that are useful as electrodes. The composite
silicon substrate electrode will generally be connected to an
electrically conductive wire to apply current and potential. The
material can be cast, coated, and/or molded onto a conductive
substrate such as a metal to form a conductive junction for
connecting conductors for transfer of current to and from the
composite electrode.
[0188] The silicon substrates of the invention generally have a
high enough electrically conductivity to act as electrodes, and to
transmit current for the oxidation and/or reduction of the bound
redox-active moieties. To make the silicon substrate more
conductive, the silicon substrate will generally include impurities
or dopants to increase electrical conductivity. Where
polycrystalline silicon is used, the polycrystalline silicon
electrode can either be deposited as doped polycrystalline silicon
(in situ doped) or can be deposited undoped and subsequently doped
with an impurity dopant such as phosphorus or boron by ion
implantation or a thermal diffusion process. Dopant impurities,
such as phosphorus and boron, tend to diffuse much more rapidly
along the grain boundaries than they do through the silicon
itself.
[0189] The dopant for a type IV semiconductor such as silicon, can
be, for example, either an electron donor or an electron acceptor.
Suitable electron donors are phosphorous, arsenic, antimony, and
bismuth. In some embodiments the dopant is phosphorous. In some
embodiments the dopant is antimony. Suitable electron acceptors are
boron, aluminum, gallium, and the like. In some embodiments, the
dopant is boron. In some embodiments, electron acceptors can impart
a chemical resistance to the silicon electrode.
[0190] Where the silicon substrate is a monolithic material such as
a wafer, the dopant can be either distributed throughout the bulk
of the silicon, or can be limited to the surface region of the
silicon wafer. In some embodiments, for example where the silicon
substrate comprises multiple zones with different redox active
moieties, the dopant can be distributed such that isolated regions
of the surface of the silicon substrate are conductive.
[0191] The dopant is generally present in an amount greater than
0.01 weight percent of the silicon, and generally in an excess of
0.1 percent of the silicon. Generally, the dopant is less than 3
weight percent of the silicon, and almost always less than 6 weight
percent of the silicon. The presence of small amounts of the dopant
can increase the electrical conductivity. The dopant may not be
homogenously distributed throughout the silicon, and the local
concentration may vary between different regions of the silicon
material.
[0192] The electrical resistivity (reciprocal of conductivity) can
be for example 0.1 (ohm-centimeters), 1 (ohm-centimeters), 10
(ohm-centimeters), 100 (ohm-centimeters), to in excess of 1000 or
even 10,000 (ohm-centimeters) or even higher which is comparable to
graphite and conventional metallic conductors.
[0193] In some embodiments the resistivity of the silicon substrate
is the range of 0.0001 to 100,000 .OMEGA.-cm. In some embodiments
the resistivity of the silicon substrate is the range of 0.001 to
10,000 .OMEGA.-cm. In some embodiments the resistivity of the
silicon substrate is the range of 0.01 to 1000 .OMEGA.-cm. In some
embodiments the resistivity of the silicon substrate is the range
of 0.1 to 100 .OMEGA.-cm. In some embodiments the resistivity of
the silicon substrate is within the range of 1 to 100 .OMEGA.-cm.
In some embodiments the resistivity of the silicon substrate is
within the range of 10 to 90 .OMEGA.-cm. In some embodiments the
silicon substrate is single crystal silicon is Si(100) that is
P-type with a resistivity of 10 to 90 .OMEGA.-cm. In some cases
several silicon substrates with different resistivities may be
used. For example, a system of the invention may comprise a one
lightly doped silicon substrate having one redox active species
bound to it, and also a more highly doped silicon surface having
another redox active species bound to it. For example, a system of
the invention may comprise one lightly doped silicon surface having
a pH sensitive redox active moiety such as anthraquinone bound
thereto, and a second silicon surface that is more highly doped
having a hydrogen ion insensitive redox active moiety such as
ferrocene bound to it.
[0194] In some cases different regions of the silicon substrate can
be doped at different levels. For example, it is well known in the
semiconductor processing art that a mask can be used to cover some
regions of the silicon, while leaving other regions exposed. The
exposed regions can be treated selectively with dopants resulting
in a silicon surface wherein some regions are doped differently
than other regions. By using multiple steps with various masks, the
conductivity properties of different regions of the silicon surface
can be controlled. Thus, some regions can have high conductivity,
some low, some regions can have a large band gap, and other regions
can have small band gaps, some regions can have N-doping, some
P-doping, and some no doping. In addition, the various regions can
be connected with conducting regions, for example deposited metal,
e.g., gold to be able to electrically address the various
regions.
[0195] In some embodiments, Nisil is used. Nisil is an alloy of
nickel and silicon. In some embodiments Nisil with 4%-5% silicon is
used. In some embodiments Nisil with 4.4% silicon is used. In some
embodiments Nicrosil is used. Nicrosil is a nickel alloy. In some
cases Nicrosil comprising 14.4% chromium, 1.4% silicon, and 0.1%
magnesium is used.
[0196] In some cases, for example, where the silicon substrate is
cast silicon, the silicon substrate will include, for example, a
silicide of a transition metal to provide castability. The silicide
of the transition metal can provide favorable mechanical properties
to the cast alloy. Typical metals useful in providing the
transition metal silicide present in the silicon electrode of the
secondary cell of this invention include titanium, zirconium,
hafnium, vanadium, columbium, chromium, molybdenum, tungsten,
manganese, iron, cobalt, nickel, copper, ruthenium, rhodium,
palladium, osmium, iridium, platinum, gold, and silver. Most
commonly, the transition metal present as the silicide in the
silicon alloy may be a silicide of manganese, chromium, iron,
cobalt, nickel, or molybdenum. The amount of the silicide may be
sufficient to provide satisfactory castability but not great enough
to deleteriously effect the properties of the silicon, i.e., from 2
percent or more up to as high as 30 percent transition metal,
elemental basis.
Redox-Active Moieties
[0197] In some embodiments, an electrochemical sensor comprises a
solid state (e.g., semiconductor) surface that is modified with
redox-active functional groups. At least one redox-active
functional group on the surface is sensitive to the presence and or
the level of a substance in the solution. In some embodiments, the
semiconductor surface can have at least one redox-active functional
group sensitive to an analyte, and at least one redox-active
functional group that is substantially insensitive to the analyte
to be tested. When used in this manner, the substantially
insensitive group can act as a reference, allowing for greater
accuracy and reproducibility of the measurements.
[0198] In some situations, an electrochemical sensor can include a
layer of a nitride, such as silicon nitride, having immobilized
thereon a redox-active moiety that is sensitive to the presence of
an analyte, such as H+, and/or a redox-active moiety that is
insensitive to the presence of the analyte.
[0199] In some situations, redox active moieties that are sensitive
and/or insensitive to the presence or absence of an analyte are
bound to a surface of a solid state working electrode, such as a
semiconductor (e.g., silicon) surface, through a surface-to-carbon
interaction, such as, e.g., a silicon-to-carbon bond in cases in
which the working electrode is formed of silicon. The
silicon-to-carbon interaction can be a covalent interaction. The
carbon atom in such a case is a carbon atom of a redox active
moiety. In other situations, redox active moieties can be bound to
a surface of a working electrode through surface-to-oxygen,
surface-to-sulfur, and/or surface-to-carbon interactions, which can
be covalent interactions.
[0200] The redox groups can be chemically or physically bound to
the surface. The redox groups can be attached to the semiconductor
covalently (e.g., via Si--C, Si--O, or Si--S bonds), can be
adsorbed to the semiconductor, or can be attached to polymers that
are either covalently or non-covalently bound to the surface.
Covalent binding of either the redox group or the polymer to which
the redox group is a part can be beneficial in improving the
lifetime and stability of the electrode. Semiconductor materials
such as silicon and germanium can form covalent bonds with carbon,
and thus is a desirable substrate for functionalizing with carbon
based molecules. The covalent binding to the surface can be through
a bond between the semiconductor, e.g., silicon, and carbon,
oxygen, nitrogen, sulfur, or other atom. In some embodiments the
bond is between silicon and carbon. In some embodiments the bond is
between silicon and oxygen. The physical bonding can occur through
adsorption, and can include, for example, spontaneous self assembly
onto the semiconductor surface of molecules such as those derived
from fatty acids which comprise redox active moieties.
[0201] Where a linker group is used, the linker can be small, for
example one to 3 atoms, or can be longer, e.g., 20 to 100 atoms, or
can be any size between large and a small linker. Where a short
linker is used, the redox-active moiety is held close to the
surface. Where a longer group is used, the redox active moiety may
be able to move away from the surface, for example into the
solution. Linker groups can comprise hydrophilic, hydrophobic
groups, or mixtures thereof. Linker groups can comprise, for
example, hydrocarbons, esters, ethers, amides, amines, carbonyls,
thiols, olefins, silicones, or other organic, inorganic or
organometallic groups. The linker groups can be formed by
polymerization or oligomerization reactions such as free radical,
cationic, or anionic polymerization. The linker group can comprise,
for example, ethylene oxide, propylene oxide, vinyl ether, or
acrylamide repeat units. Linkers can have ring structures including
aromatic rings. The variation in the linker structure can be used
to vary the mobility of the redox-active moiety in the
solution.
[0202] As used herein, the term moiety generally refers to a
portion of a molecule or substituent. A redox-active moiety may be
highly substituted, and can still act as a redox-active moiety. As
used herein, the terms "redox active moiety", "redox active group",
"redox active functional group", and "redox group" are used
interchangeably. Thus, for example, the redox-active moiety
ferrocene includes substituted ferrocenes, ferrocene polymers, and
ferrocene covalently attached to the surface with or without linker
molecules.
[0203] In some embodiments, the redox-active moiety can be
incorporated into a polymer, and the polymer comprising the redox
active moiety can be immobilized onto the semiconductor surface.
The immobilization of the polymer can be either chemical or
physical. The immobilization of the polymer can be through covalent
bonds, or through adsorption of the polymer to the semiconductor
surface.
[0204] In some embodiments, the redox-active moiety is bound to a
particle that is bound to semiconductor. The particle is generally
an electrically conductive particle. The particle attached to the
semiconductor surface in a manner that allow for current to flow
between the semiconductor surface and the particle. The particles
can be attached chemically or physically to the surface. For
example, the redox-active moiety can be attached to a carbon
particle, and the carbon particle attached to the semiconductor
surface. In some embodiments, the carbon particle can be a carbon
nanotube. In some embodiments of the invention, carbon nanotubes
can be attached to the surface of the semiconductor, where there
are redox-active groups attached to the carbon nanotubes. For
instance, attachment of well-aligned single-walled carbon nanotubes
architecture to a single-crystal silicon surface can be used. In
some embodiments, for example, ferrocenemethanol molecules are
attached to single walled carbon nanotube (SWCNT) arrays that are
directly anchored to the silicon surface, for example, a (100)
surface. For example, single wall carbon nanotubes can be coupled
to the surface using this method as described in Yu et al.,
Electrochimica Acta 52 (2007) 6206-6211.
[0205] The redox-active moieties generally have reversible redox
activity with well-defined cyclic voltammetry oxidation and/or
reduction peaks. A suitable reference redox reagent can vary from
application to application or medium to medium depending on the
intended use.
[0206] The position of the reduction and/or oxidation potentials of
the redox active moiety can be chosen to improve the accuracy and
quality of the measurement of redox potential. In some cases, the
reduction and or oxidation potential can be chosen to be away from
other redox active species. The silicon surface, for example,
generally has a wide window in which to perform measurement of
reduction or oxidation potential without interfering with the
measurement of the reduction and or oxidation of the redox active
moieties bound to the surface. The silicon surface can generally be
used to measure oxidation and/or reduction potentials from between
negative 2 V to positive 2 V. In some cases, for example where the
medium is an aqueous medium, the reduction and/or oxidation
potential of the redox-active moiety can be chosen so as not to
fall within the reduction or oxidation potential of the medium to
minimize interference. This can be useful where cyclic voltammetry
is used, and is less important when square wave voltammetry is
used.
[0207] Redox-active moieties that are insensitive to the presence
of analytes should show little or no change in their oxidation
and/or reduction potentials in the presence or absence of such
analytes. Redox-active moieties that are generally insensitive to
the presence of analytes, and in particular are insensitive to the
presence of hydrogen ion include: ferrocene, polyvinylferrocene,
Os(bpy).sub.2Cl.sub.2, Ru(bpy).sub.2Cl.sub.2, viologen,
polyviologen, and polythiophene. Redox reagents having a high
degree of electrochemical reversibility are generally preferred.
FIG. 5 shows examples of the hydrogen ion insensitive redox-active
moiety ferrocene bound to a silicon surface.
[0208] Non-limiting redox-active moieties that are sensitive to
hydrogen ion include: quinones, anthroquinones, phenanthroquinones,
phenylene diamines, catechols, phenothiazinium dyes, and
monoquaternized N-alkyl-4,4'-bipyridinium. In some embodiments the
redox-active moiety that is sensitive to the presence of hydrogen
ion can include inorganic materials and metal oxides. Hydrogen ion
sensitive inorganic redox-active inorganic moieties include
Prussian Blue, Ni(OH).sub.2, and RuO.sub.x. FIG. 6 shows examples
of the hydrogen ion sensitive redox-active moiety anthracene
covalently bound to a silicon surface. FIG. 7 shows an example of a
silicon surface having covalently bound thereto both the hydrogen
ion sensitive redox-active moiety ferrocene and the hydrogen ion
insensitive redox-active agent anthracene.
[0209] In some embodiments the analyte is carbon monoxide (CO). An
example of a CO sensitive redox-active agent is ferrocenyl
ferrazetine disulfide or ferrazetine-ferrazetine disulfide. A CO
insensitive redox-active agent can be, for example, ferrocene.
[0210] In some embodiments the analyte is an alkali metal. Alkali
metal sensitive redox-active agents include, for example:
1,1'-(1,4,10,13-tetraoxa-7,16-diazacyclooctadecane-7,16-diyl
dimethyl), ferrocenyl thiol, and other ferrocene derivatives
containing covalently attached cryptands. These materials are
described, for example, Hammond, et al., J. Chem. Soc. Perkin.
Trans. 1707 (1983); Medina, et al., J. Chem. Soc. Chem. Commun. 290
(1991); Shu and Wrighton, J. Phys. Chem. 92, 5221 (1988). Included
are examples such as the above ferrocenyl ferrazetine and
ferrocenyl cryptand, in which an ordinarily chemically insensitive
redox center (ferrocene) is covalently linked to a chemical
recognition site in such a way as to make the redox center
chemically sensitive. Also suitable are molecules or polymers in
which the sensor and reference functionalities are covalently
linked such as 1-hydro-1'-(6-(pyrrol-1-yl)hexyl)-4,4'-bipyridinium
bis(hexafluorophosphate), as described by Shu and Wrighton, J.
Phys. Chem. 92, 5221 (1988).
[0211] In some cases, more than one analyte-sensitive redox-active
moiety can be used. For example, there can be one redox-active
moiety that is sensitive to hydrogen ion, and another that is
sensitive to a second analyte such as CO, oxygen, ammonia, or an
alkali metal. This approach allows for the simultaneous measurement
of several analytes. In some cases redox-active agents sensitive to
3 or more analytes can be used. Where there are redox agents
sensitive to multiple analytes, in many cases, it is also desired
to have one or more analyte-insensitive redox-active moieties bound
to the semiconductor surface as well to provide a reference, to
improve accuracy, and to minimize or avoid calibration.
[0212] Where more than one redox-active moiety is used it can be
important to ensure there is not significant interference between
the peaks. This can be desirable especially when the multiple
redox-active moieties are on the same electrically addressable zone
on the semiconductor substrate. This can be achieved, for example,
by ensuring that there is sufficient separation of the oxidation
and reduction potentials, or by physically separating the redox
moieties.
[0213] In some embodiments, the semiconductor substrate has a
plurality of isolated separately electrically addressable zones. In
some embodiments, the different zones will comprise different
redox-active moieties. The use of separate zones can be beneficial
in that the voltammetric measurements can be carried out
separately, allowing for the use of multiple redox-active agents
with that have similar reduction and/or oxidation potentials.
[0214] The separately electrically addressable zones can be made by
conventional semiconductor processing methods, for example masking
to create structures in specific areas on the surface, for example
specific areas on the surface having certain levels of doping.
Conventional semiconductor processing can also be used to
incorporate conductive interconnects allowing the zones to be
separately addressable. Masking can also be used during the
attachment of the redox-active moieties to the semiconductor
surface to attach specific redox active moieties to different
regions of the surface.
[0215] The semiconductor substrate with separately addressable
zones can effectively create an electrochemical sensor array.
Another aspect of the invention provides a semiconductor
electrochemical sensor array wherein a plurality of zones, each
zone comprising a redox active moiety. The array can have multiple
zones with analyte-sensitive redox-active moieties, and one or more
zones with analyte-insensitive redox active moieties. A zone can
have a single redox-active moiety, or multiple redox active
moieties. The array can be constructed, for example, to measure
both pH and O.sub.2, wherein one zone comprises a redox-active
moiety sensitive to hydrogen ion, another zone has a redox active
moiety sensitive to O.sub.2, and a third zone with a redox-active
moiety that is insensitive to both hydrogen ion and O.sub.2.
[0216] In some embodiments, an array of separately addressable
zones or an array of electrodes is used where there are a plurality
of zones or electrodes that are each constructed to measure the
same analyte and used in a redundant matter, wherein another zone
capable of measuring the same analyte is used either simultaneously
or in place of the other zone or electrode. In some cases, more
than one zone or electrode, for example in an array, is used
simultaneously to improve the quality of the measurement. In some
cases, more than one zone or electrode, for example in an array, is
used sequentially, wherein if one zone or electrode shows degraded
performance, the measurement of that analyte is performed on
another zone or electrode constructed to have similar
characteristics. The sequential use of similar zones or electrodes
can provide reliability of measurement over time. While the
electrodes of the present invention can be prepared to be robust
and to resist fouling, in some circumstances, degradation of the
measurement over time may occur. In some embodiments, the maximum
current (I.sub.max) can be monitored over time. A system can be
configured, for example, such that when the maximum current drops
below a certain level, a switch is made to a redundant zone or
electrode for further measurement of that analyte. There can be
multiple redundant elements, e.g., 1, 2, 3, 4, 5, 10, 20 or more
redundant elements.
[0217] The semiconductor substrate of the invention can also
comprise circuitry. The circuitry can be used, for example for
controlling the current and potential provide to the redox-active
moiety. The circuitry can also be used for analyzing signals or for
processing data related to the voltammetric measurement. The
circuitry can also have other functionality, such as the ability to
measure other parameters such as temperature, the ability to store
date, or the ability to send data and receive instructions from a
remote location. The circuitry can contain, for example, an
amplifier such as an operational amplifier. The circuitry can
contain, for example, an analog to digital converter (ADC). In some
cases having the amplifying and ADC functions incorporated into the
semiconductor substrate can provider higher quality and reliability
of the transmission of the signal from the sensor. The use of
circuitry on the semiconductor substrate can be particularly useful
when an array of zones is utilized. The circuitry can assist in
managing the passage of current in and out of the semiconductor
substrate. In some cases, the circuits can allow for schemes to
simultaneously or sequentially address the zones, for example, by
mutiplexing (MUX).
Solid State Electrode Sensors
[0218] Another aspect of the invention provides a solid state
sensor having a solid state working electrode formed of a solid
state (e.g., semiconductor) substrate. The solid state electrode
sensor comprising the solid state substrate can be used for the
measurement of the presence or absence of one or more analytes, or
can be used to accurately measure the concentration of analyte in a
sample.
[0219] In some embodiments, a solid state electrode sensor is
formed of a semiconductor, such as silicon. The semiconductor
electrode sensor can be used to detect the presence or absence
and/or the measure the concentration of analytes, including
hydrogen ion, alkali metals, CO, or O.sub.2. In some embodiments,
the semiconductor electrode sensor is used to measure the
concentration of hydrogen ion, or pH.
[0220] The semiconductor electrode sensor comprises a semiconductor
substrate as described above comprising a redox-active moiety that
is sensitive to the presence of an analyte. The semiconductor
electrode sensor may also comprise a redox active moiety that is
insensitive to the presence of an analyte. The semiconductor
electrode sensor can comprise more than one semiconductor
substrate. For example, the sensor may comprise one semiconductor
substrate that has an analyte sensitive redox active moiety and
another semiconductor substrate having an analyte insensitive redox
active moiety.
[0221] The semiconductor electrode sensor is configured to be
incorporated into a system that will supply voltage, and can drive
current to the sensor to perform voltammetry. Thus, the
semiconductor substrate or substrates within the sensor may be
electrically connected in a manner which will allow for connection
to a device for supplying and measuring current and voltage.
[0222] The semiconductor electrode sensor is generally the working
electrode in an electrochemical system that will also comprise a
counter electrode, and in some embodiments, a reference
electrode.
[0223] The sensor may be put into contact with a sample having the
analyte to be detected. The sample is generally a liquid sample. In
some cases the sample can be a gel, suspension, molten, or
semi-solid medium. The sample can be, for example, a foodstuff. The
sample can be any type of liquid including hydrocarbons, oils,
fluorocarbons, silicones, and aqueous solutions. Where the analyte
is hydrogen ion, an aqueous medium is generally used, but in some
case a polar protic medium or polar aprotic medium can be used. The
sensor is useful for measuring pH in aqueous solutions.
[0224] In some embodiments, the sensor of the invention can
accurately measure analyte concentrations where the analyte is
present in a concentration range from 10.sup.-1M to 10.sup.-14M. In
some embodiments, the sensor of the invention can accurately
measure analyte concentrations where the analyte is present in a
concentration range from 10.sup.-3 M to 10.sup.-10 M. In some
embodiments, the sensor of the invention can measure the
concentration to an accuracy of plus or minus 100%, 50%, 30%, 20%,
10%, 5%, 2% or 1%. In some embodiments, the sensor of the invention
can measure the concentration within a range of 10.sup.-3 M to
10.sup.-10 M to an accuracy of plus or minus 100%, 50%, 30%, 20%,
10%, 5%, 2% or 1%.
[0225] In some embodiments, the analyte is hydrogen ion, and the
sensor of the invention can accurately measure the pH in a range
from pH 1 to pH 14. In some embodiments, the analyte is hydrogen
ion, and the sensor of the invention can accurately measure the pH
in a range from pH 3 to pH 10. In some embodiments, the sensor of
the invention can accurately measure pH to an accuracy of plus or
minus 0.5, 0.3, 0.2, 0.1, 0.07, 0.05, 0.03, 0.02, or 0.01 pH units.
In some embodiments, the sensor of the invention can accurately
measure pH in a range from pH 3 to pH 10 to an accuracy of plus or
minus 0.5, 0.3, 0.2, 0.1, 0.07, 0.05, 0.03, 0.02, or 0.01 pH
units.
[0226] The semiconductor electrode sensors of the invention can
accurately measure analyte concentration in a wide variety of
sample types. The sensors can be made to be robust, and resistant
to fouling, and therefore reliable for long-term measurements.
[0227] Another aspect of the invention is a sensor which does not
require routine calibration (or re-calibration), or in some cases
any calibration at all. Conventional potentiometric sensors rely on
a glass membrane to sense, for example, hydrogen ion. These types
of sensors generally need to be calibrated on a regular basis,
usually by placing the sensor into standards of known analyte
concentration. These types of sensors generally need calibration
when going from one solution to another solution, and will also
need calibration with time, even if kept within the same solution
and upon standing outside of a solution. The situation is made
worse if there is a change in the composition of the medium over
the time that the sensor is monitoring the medium, for example,
when monitoring a chemical reaction, biochemical reaction, or
fermentation. In these cases, potentiometric sensors may drift and
need calibration due to the accumulation of some species in the
reaction, or due to fouling of the sensor by species present.
[0228] In some embodiments, the sensors of the present invention do
not need to be calibrated under any of these situations. In some
embodiments, the sensors of the invention do not need to be
calibrated over time in solution. In some embodiments, the sensors
of the invention do not need to be calibrated after an hour, 10
hours, 1 day, 2 days, 5 days, a week, two weeks, a month, 6 months,
1 year, 2 years or longer while in a solution or in storage. In
some embodiments the sensors or the present invention are accurate
at measuring analyte concentration to 50%, 40%, 20%, 10%, 5%, 2%,
1%, 0.5%, 0.2% or 0.1% after the times above. In some embodiments
where the sensors measure pH, the sensors are accurate to 1, 0.8,
0.5, 0.3, 0.2, 0.1, 0.08, 0.05, 0.03, 0.02, or 0.01 pH units after
the times above. In some embodiments where the sensors measure pH,
the sensors are accurate to within 0.1 pH units after one week in
solution or in storage.
[0229] In some embodiments, the sensor is capable of measuring
analyte concentration without any calibration with an external
standard. In some embodiments, the sensor remains sensitive to the
analyte without calibration after a first use by an end user.
[0230] In some embodiments the analyte is hydrogen ion and the
sensor remains sensitive to hydrogen ion after exposure to a cell
culture medium for at least 1, 3, 6, 9, 12, 18, or 24 hours or 2,
3, 4, 6, 8, 12, 24, 48, 60, 90, or more days. In some embodiments
the analyte is hydrogen ion and the sensor remains sensitive to
hydrogen ion after exposure to a cell culture medium for at least 3
days. In some embodiments the analyte is hydrogen ion and the
sensor remains sensitive to hydrogen ion after exposure to a cell
culture medium for at least 6 days. In some embodiments, the sensor
is capable of measuring pH with an accuracy of 0.2 units after
exposure to the cell culture medium.
[0231] In some embodiments, the analyte is hydrogen ion and the
sensor is capable of measuring pH with an accuracy of 0.2 units
after autoclave treatment at 121.degree. C. for 10, 20, 40, 80,
100, 200, 400, or 800 minutes. In some embodiments, the analyte is
hydrogen ion and the sensor is capable of measuring pH with an
accuracy of 0.2 units after autoclave treatment at 121.degree. C.
for 40 minutes. In some embodiments, the analyte is hydrogen ion
and the sensor is capable of measuring pH with an accuracy of 0.2
units after autoclave treatment at 121.degree. C. for 400
minutes.
[0232] A subject sensor that does not require calibration over long
periods of time in a medium that can change characteristics is
useful, for example, as an implantable sensor. The implantable
sensor can be placed under the skin or within the body in contact
with a bodily fluid such as blood, saliva, breast milk, amniotic
fluid, lymph, sweat, tears, or urine. The sensor can measure the
concentration of analytes such as hydrogen ion, sodium, potassium,
calcium, or oxygen.
[0233] The implantable sensor has an electrode configured to be in
contact with a bodily fluid, said electrode comprising a
semiconductor surface that has immobilized thereon a redox active
moiety, wherein the redox active moiety has an oxidation potential
and/or reduction potential that is sensitive to concentration of
said ion. The implantable sensor can be included in an implantable
medical device such as described in U.S. Pat. No. 6,738,670. For
example, the implantable medical device in which the sensor resides
could include pacemakers, defibrillators, drug delivery pumps,
diagnostic recorders, cochlear implants, and the like. The
implantable medical device is typically programmed with a therapy
and then implanted in the body typically in a subcutaneous pocket
at a site selected after considering clinician and patient
preferences. In some embodiments the implanted device is in a form
which can be swallowed, allowing the measurement of the properties
of the regions encountered as it passes through the body such as
the digestive tract including the stomach, the upper and lower
intestines, and the colon. The information obtained by the sensor
in the implanted device can either be accessed in real time, for
example, by wireless communication, or can be retrieved from the
device after passage through the body. A wide variety of
programmers, also known as downlink transmitters, can be used to
transmit data to and receive data from the implantable medical
device. Examples of downlink transmitters include devices such as
physician programmers, patient programmers, programming wands,
telemetry access units, and the like. The clinician, for example,
can periodically use a physician programmer to communicate with the
implantable medical device to manage the patient's therapy and
collect implantable medical device data. The semiconductor
electrode sensor can be incorporated into or attached to the
implantable medical device and can provide data on analyte
concentration within the region of the body into which it is
implanted. The patient can use the patient programmer to
communicate with the implanted device to make therapy adjustments
that have been programmed by the clinician. Both the physician
programmer and patient programmer can have an antenna locator that
indicates when a telemetry head is aligned closely enough with the
implanted device for adequate telemetry.
[0234] Another aspect of the invention is a method for measuring
concentration in a bodily fluid within a body, the method
comprising placing a semiconductor electrode sensor comprising a
redox active moiety in contact with the bodily fluid, and operating
the sensor to yield a value of the concentration of the analyte
present in said bodily fluid.
Systems for Measuring the Concentration and/or Presence or Absence
of an Analyte
[0235] Another aspect of the invention provides a system for
measuring analyte concentration. In some embodiments, the system
comprises a working electrode having a solid state (e.g.,
semiconductor) surface that has immobilized thereon a redox active
moiety, wherein the redox active moiety has an oxidation potential
and/or reduction potential that is sensitive to the presence of an
analyte; a counter electrode and optionally a reference electrode.
The system further comprises a source for supplying a plurality of
potentials to the working electrode, and a device for measuring
current through the working electrode at the plurality of
potentials. The working electrode referred to herein can comprise
the solid state electrochemical sensor described above. It is
desirable in some embodiments that the solid state surface also has
immobilized thereon a second redox active moiety having an
oxidation potential and/or reduction potential that is insensitive
to the presence of said analyte. The redox active moiety that is
insensitive to the presence of the analyte can be on the same solid
state surface, or can be on another solid state surface in
electrical communication with the system and in contact with the
sample. In some embodiments, the redox active moiety that is
sensitive to the presence of the analyte is on first solid state
(e.g., semiconductor) working electrode, and the redox active
moiety that is insensitive to the presence of the analyte is on a
second solid state working electrode that is electrically isolated
(or electrically insulated) from the first working electrode. The
system is configured such that the working electrode(s), the
counter electrode, and optionally the reference electrode are in
contact with the sample. In many embodiments, the sample is a
liquid sample, and the electrodes are each in contact with the
liquid. In some cases, the sample will not be a liquid, but may be
a solid, generally comprising a solid electrolyte, a semi-solid
(e.g., solid-liquid mixture), or a gas, or a sample having a
viscosity characteristic of a gas or liquid. In some embodiments,
the first solid state working electrode is separately and
independently addressable from the second solid state working
electrode, enabling a reading from the first working electrode
independently from the second working electrode.
[0236] In some embodiments, the system will have two or more
working electrodes. For example, in some embodiments, the system
will have one working electrode comprising a semiconductor surface
that has immobilized thereon a redox active moiety whose oxidation
potential and/or reduction potential is sensitive to the presence
of said analyte, and a second working electrode comprising redox
active moiety whose oxidation potential and/or reduction potential
is insensitive to the presence of said analyte. An example of a
system with two working electrodes is a system having two
semiconductor wafers or chips, one of which has a redox active
moiety which is sensitive to pH, such as anthracene, and another
redox active moiety which is insensitive to pH, such as a
ferrocene. In some cases the semiconductor wafer or chip on which
each redox active species is immobilized may be a different type of
semiconductor wafer or chip. For instance, the semiconductor wafer
or chip to which the pH sensitive moiety is bound may have one
doping level, and the semiconductor wafer or chip on which the pH
insensitive moiety is bound may have a different doping level. This
type of construction can be beneficial because, in some cases, one
type of redox active species will perform better in terms of
amplitude, sensitivity or stability with one type of doping, while
another redox active species will perform better on a semiconductor
wafer or chip with a different type of doping. In some embodiments,
the pH sensitive moiety, e.g., anthracene, is bound to a
semiconductor wafer that has a low level of doping, and the pH
insensitive moiety, e.g., ferrocene is bound to a silicon wafer
that has a higher level of doping. In some embodiments the
semiconductor wafer onto which the pH sensitive moiety, e.g.,
anthracene is bound has a resistivity between 1 .OMEGA.-cm an 1000
.OMEGA.-cm, or between 10 .OMEGA.-cm and 90 .OMEGA.-cm, or between
10 .OMEGA.-cm and 40 .OMEGA.-cm, while semiconductor wafer onto
which the pH insensitive moiety, e.g., ferrocene, is bound has a
resistivity between 1 milliohm-cm and 1000 .OMEGA.-cm, or 1
.OMEGA.-cm and 90 .OMEGA.-cm, or 10 .OMEGA.-cm and 40 .OMEGA.-cm.
In some embodiments, an N-type semiconductor (e.g., silicon wafer)
is used for a working electrode having an H+ insensitive moiety
(e.g., ferrocene). In some situations, an undoped or lightly doped
p-type semiconductor (e.g., silicon wafer) is used for a working
electrode having an H+ sensitive moiety (e.g., anthracene). In
other embodiments, a P-type semiconductor (e.g., silicon wafer) may
be used for both the pH sensitive and the pH insensitive
moieties.
[0237] In some embodiments, the system will have 3 or more working
electrodes. For example, in some embodiments, the system will have
one working electrode comprising a semiconductor surface that has
immobilized thereon a redox active moiety that is sensitive to the
presence of a first analyte, a second working electrode comprising
a semiconductor surface that has immobilized thereon a redox active
moiety that is sensitive to the presence of a second analyte, and a
third working electrode comprising a semiconductor surface that has
immobilized thereon a redox active moiety that is insensitive to
the presence of either the first analyte nor the second analyte.
The system can also have more than 3 working electrodes, for
example having 4, 5, 6, 7, 8, 9, 10, 12, 20, 50 or more working
electrodes, each having redox active moieties sensitive to and
analyte. These systems can also have one or more than one
semiconductor working electrode having a redox species that is
insensitive to the analytes, for example to provide a reference. In
some cases more than one redox species that is insensitive to the
analyte can be used.
[0238] In some embodiments the system of the present invention
comprises a probe that comprises the 2 or more electrodes. The
probe can physically hold the electrodes such that the electrodes
can be brought into contact with the sample. The probe allows the
working electrodes to be held close to the reference and/or counter
electrode. FIG. 21 shows an exemplary embodiment of a probe of a
system of the present invention having 4 electrodes: a first
working electrode (WE1), a second working electrode (WE2), a
reference electrode (RE), and a counter electrode (CE). WE1 and WE2
can each be formed of a solid state material, such as a
semiconductor. FIG. 21(a) shows a shaded drawing of the probe. FIG.
21(b) shows a line drawing of the probe. As shown in FIG. 21, in
some cases it can be beneficial in the present invention for the
reference electrode and the counter electrode to have a ring
configuration. In other embodiments, only one of the reference
electrode or counter electrode will have a ring configuration. A
ring electrode can in some cases provide signal stability. While
this embodiment shows one configuration, there are other ring
electrode configurations that can be used with the present
invention.
[0239] In some embodiments it is useful to use undoped or lightly
doped semiconductor (e.g., silicon) substrates for a moiety such as
anthracene. While not being bound by theory, the band gap of the
semiconductor, e.g., silicon can be influenced by the level of
doping of the semiconductor, and it is believed that in some cases,
the use of a semiconductor with the appropriate level of doping can
be useful to tailor the appropriate redox active moiety with the
appropriate band gap. Thus in some cases it is desirable to use
lightly doped semiconductor, e.g., silicon with a moiety such as
anthracene. Thus in some embodiments it is useful to use two
semiconductor working electrodes: one optimized for ferrocene
moieties and the other optimized for the anthracene moieties. When
two working electrodes are used, in some embodiments, two
sequential electrochemical measurements (e.g., with square wave
voltammetry) may be carried out using the same counter and
reference electrode. For instance, the first measurement can be
conducted using reference, counter and working electrode 1 (e.g.,
anthracene derivatized) between -1.2 to -0.5 V, followed by the
second measurement which may be conducted using reference, counter
and working electrode 2 (e.g., ferrocene derivatized) between 0 to
0.5 V. The peaks potential detected in the first and second
measurements can then be stored and processed to get a pH reading.
This type of system and method can be accomplished through the use
of a bipotentiostat or a two-channel multiplexer. A similar
approach can be applied to multiple working electrodes with a
mutipotentiostat or a multi-channel multiplexer.
[0240] The system is configured to carry out voltammetric
measurements on the sample. Some embodiments provide a method which
includes the measurement of pH with a voltammetric pH sensing
system comprising the semiconductor electrode sensor described
above, a potentiostat for providing voltage to the electrodes, and
a meter for detecting the current as a function of voltage.
[0241] The counter electrode typically is needed to complete the
electrochemical circuit to make the measurements described herein.
The counter electrode is generally made of a material which is
electrochemically inert to the medium so that current overloading
does not occur during the course of measurement. Suitable materials
in many applications include platinum, silver, gold, stainless
steel, and carbon.
[0242] A reference electrode is optional and is used as a third
electrode in some embodiments of the invention. In the case of a
three-electrode system, the counter electrode generally completes
the circuit, allowing current to flow through the cell, while the
reference electrode maintains a constant interfacial potential
difference regardless of the current. In the case where the system
comprises an analyte sensitive redox active moiety and an analyte
insensitive redox active moiety, the analyte insensitive redox
active moiety can act as a reference, allowing the potential
difference to be used to determine analyte concentration. Even
where the system comprises an analyte insensitive moiety, in some
embodiments, a reference electrode will still be used. In some
embodiments, pseudo-reference electrodes can also be utilized.
Reference electrodes that can be employed include: Standard
hydrogen electrode (SHE), also known as "normal hydrogen electrode"
(NHE), saturated calomel electrode (SCE), copper-copper(II) sulfate
electrode, and silver/silver chloride (Ag/AgCl) electrode. In some
embodiments a silver electrode or a polyurethane coated silver
electrode can act as a silver/silver chloride electrode where
sufficient chloride is present at or near the silver electrode. In
some cases, e.g., in non-aqueous medium, a metal electrode such as
a platinum or a silver electrode, or a polyurethane coated silver
electrode can be used as the reference electrode.
[0243] To carry out voltammetry, the system generally has a source
for supplying a plurality of potentials. The voltammetry can be,
for example cyclic voltammetry, pulse voltammetry, normal pulse
voltammetry, square wave voltammetry, differential pulse
voltammetry, linear voltammetry, or square wave voltammetry. The
source for supplying a plurality of potentials can be a
potentiostat, for example, a potentiostat capable of applying
square waves for square wave voltammetry.
[0244] Generally, the analyte concentration is determined by using
voltammetry to identify the position of current peaks, which
current peaks indicate the reduction or oxidation potential of a
redox active moiety. In some embodiments, the position of the
reduction and/or oxidation potential of the analyte sensitive redox
active moiety is used to determine the concentration of the
analyte. This method can be used, for example, where no analyte
insensitive redox active moiety is employed.
[0245] Where an analyte insensitive redox active moiety is used,
detection is generally accomplished by measuring the potential
difference, delta E, associated with current peaks for oxidation
(or reduction) of the immobilized redox active moieties, where the
magnitude of delta E can be related to the concentration of
analyte, e.g., hydrogen ion (H+) in solution. The analyte
insensitive redox active moiety has an electrochemical response
that is insensitive to variations in the medium and serves as the
reference. Current peaks for oxidation or reduction of the
reference and indicator are determined from a voltammogram using a
counter electrode.
[0246] In some embodiments, the system further comprises a
computation system that communicates with the device for measuring
current. The computation system can have algorithms for calculating
reduction or oxidation potential from the measured current at a
plurality of potentials from the voltammetry measurements. The
computing systems can be part of the sensing system, in some cases
allowing the sensing system to be self-contained. The computing
system can comprise memory for storing raw and/or processed data
from the sensors. The computing system can be connected to a
transmission device that will wirelessly, by wire, fiber or other
means, transmit processed data to an external device. The computing
system can provide signals and measurements which can be
transmitted in some cases in real time, allowing the system to
alert end users of conditions which may require attention. The
transmitted signals and measurements can, for example, provide the
information required for adjusting a manufacturing process such as
a chemical or biochemical process.
[0247] In some embodiments the system is made up of a housing that
holds the semiconductor electrode sensor that is electrically
connected to a unit comprising the source for supplying a plurality
of potentials and the current measuring device. In some embodiments
the unit also comprises the computing system described above for at
least partially analyzing the data. The unit can be self powered,
e.g., with a battery, or can have a connection to an outside power
source. The unit can have a display and input buttons to allow the
user to control the measurement and to read the output from the
sensor. The unit can have transmission capability for sending out
data, and for receiving instructions or to be tested by an external
device.
[0248] FIG. 1 shows drawings of an embodiment of the connection of
a semiconductor sensor electrode into a sensor housing for use in
measuring analytes in a fluid such as the fluid in a biochemical
reactor. FIG. 1(a) shows a blow up drawing of an assembly that
holds the semiconductor electrode sensor and provides electrical
connections to the semiconductor electrode sensor for voltammetry
measurements. In FIG. 1(a) the semiconductor electrode sensor (I),
is held in place by the end cap (II), in contact with the
metallized ceramic disk (III). The ceramic disk can be metallized
in specific areas on both the front and the back of the disk, with
vias connecting the specific metallized areas. On one side of the
ceramic disk, the semiconductor working electrodes as well as the
counter and optionally reference electrodes can be present. For
example, electrodes such as the semiconductor working electrodes
can each being mounted to a specific metallized area. The disk is
then mounted into the housing such that the side of the disk having
the electrodes is exposed to the medium into which the probe is
immersed, and the other side of the disk is away from the medium,
allowing for electrical connection to the metallized areas on to
the disk such that voltage can be applied and current can flow to
and from the electrodes. The sealing gasket (IV) provides sealing
from the fluid in which the sensor is immersed while allowing
electrical contact with the pin contacts (V) on the shaft of the
housing (VI). FIG. 1(b) shows the housing assembled for insertion
into the fluid to be measured. Pipe fittings are used to seal the
wires that provide electrical connection to the semiconductor
electrode for voltammetric measurements.
[0249] FIG. 2 shows a drawing of an embodiment of the unit
comprising the source for supplying a plurality of potentials and
the current measuring device. FIG. 2(a) shows a top view and FIG.
2(b) shows a back side view. The unit has an electrical
input/output connector for connecting to the electrodes for
carrying out voltammetry. The unit has a connection for AC power.
The unit has a universal serial bus interface (USB I/F) and an
RS-232 Serial port for transmitting data, for receiving
instructions, and for testing and debugging by an external device.
The unit also comprises a liquid crystal display (LCD) and has user
interface buttons to allow the user to control the measurements and
to read the output from the sensor.
[0250] FIG. 3 shows another exemplary embodiment of a system, in
accordance with an embodiment of the invention. FIG. 3 shows a
probe that contains two semiconductor (silicon) working electrodes,
a reference electrode, and a counter electrode. The electrodes can
be electrically connected through the probe to the source for
supplying a plurality of potentials to the working electrode, the
counter electrode, and optionally the reference electrode, and a
device for measuring current through the working electrode at the
plurality of potentials. In this embodiment, the two working
electrodes, the reference electrodes and the counter electrode are
contained on the end of the probe on a disk which allows the
electrodes to be in contact with the medium comprising the
analyte(s) to be measured. The areas of the various electrodes can
be varied to improve the performance of the system. While this
embodiment shows two working electrodes, in some embodiments, there
may be one working electrode, and in other embodiments, there are
3, 4, 5, 10, 20, or more working electrodes. In some embodiments
one working electrode can comprise a semiconductor surface with a
redox active moiety that is sensitive to pH, such as anthracene,
and the other working electrode can comprise a redox active moiety
that is insensitive to pH, such as ferrocene. In some embodiments,
the semiconductor surface on which the pH sensitive moiety, such as
anthracene, is bound comprises a silicon wafer that is lightly
doped, and the silicon surface on which the pH insensitive moiety,
such as ferrocene, is bound comprises a silicon wafer that is more
heavily doped.
[0251] The semiconductor working electrodes can be bonded to
conductive regions on the disk. Conductive vias through the disk
allow electrical connection to electrode pins which are contained
within the housing. The electrode pins are in turn, electrically
connected, for example to wires, which can run through the probe
housing, and then out of the housing to electrically connect the
electrodes to the source for applying potentials to the electrode,
and to the device for measuring the current that passes through the
electrodes. The threads, o-ring, and hex body allow for the probe
to be mounted into a reactor such as a bioreactor or fermentor. The
threads allow for the probe to mate with a corresponding threaded
hole through the wall of the reactor, the hex body allows for
tightening the probe into the reactor, and the gasket assists in
establishing a seal. In some embodiments the unit is mounted into
the reactor without the use of threads, mounting without threads
can provide resistance to some failure modes that are present when
threads are used.
[0252] In some embodiments, the system is configured to be used as
an in-line sensor in a process. An in-line sensor can be a sensor
that is used in an on-going process. In some embodiments the sensor
is in a vessel, in other embodiments the sensor is in a conduit or
pipe through which a process fluid flows. In some embodiments, the
currents measured at a plurality of potentials by voltammetry are
used to determine analyte concentration, and the determined analyte
concentration is used to control a process parameter. The systems
of the present invention are valuable in in-line sensing in that
they can be made to be robust, to resist fouling, and are able to
measure analyte concentration for long periods of time in media
that changes its properties, as in a process such as a chemical
reaction.
Methods for Forming Solid State Electrochemical Sensors
[0253] Another aspect of the invention provides a method for
forming a solid state electrochemical sensor. The method generally
comprises having a solid state (e.g., semiconductor) substrate with
a surface and immobilizing a redox active moiety with a reduction
and/or oxidation potential that is sensitive to an analyte onto the
solid state surface.
[0254] Some embodiments provide a method for forming an
analyte-sensitive semiconductor electrode, the electrode having a
semiconductor surface. The method comprises immobilizing a
redox-active moiety that is sensitive to the presence of an analyte
onto the semiconductor surface.
[0255] Any suitable method such as those known in the art or
disclosed herein can be used to construct a semiconductor surface
as described above that is useful as part of a subject sensor. The
redox groups can be immobilized onto the surface chemically or
physically. The redox groups can be reacted with the semiconductor
surface to attach them to the semiconductor covalently.
Alternatively, the redox active groups can be adsorbed to the
semiconductor. The redox active groups can also be immobilized by
attaching the groups to a polymer that is either covalently or
non-covalently bound to the surface. Covalent binding of either the
redox group or the polymer to which the redox group is a part to
the surface can be beneficial in improving the lifetime and
stability of the electrode.
[0256] Functional groups can be covalently attached to
semiconductors, such as silicon or germanium. Silicon can, for
example, form covalent bonds with carbon, and thus is a desirable
substrate for functionalizing with carbon based molecules. The
covalent binding to the surface can be through a bond between the
semiconductor, e.g., silicon, and carbon, oxygen, nitrogen, sulfur,
or other atom. In some embodiments the bond to the surface is
between the semiconductor, e.g., silicon, and carbon. In some
embodiments the bond to the surface is between semiconductor, e.g.,
silicon, and oxygen.
[0257] In some embodiments, the immobilization of the redox active
moiety by covalent binding to the semiconductor surface is
accomplished by reaction with a semiconductor hydride, for example,
a silicon hydride (Si--H) surface. A semiconductor-hydride, e.g.,
silicon-hydride surface can be obtained, for example by treatment
of the semiconductor, e.g., silicon surface, for example a surface
that is in the native oxide state, with hydrofluoric acid (HF). For
example, dilute (1-3%) aqueous HF treatment, or, a 40% aqueous
NH.sub.4F treatment can be used to create a Si--H terminated
surface. Porous silicon, when etched through standard procedures
involving HF, can also be used as a Si--H surface. FIG. 4 shows a
schematic illustrating the conversion of a native oxide surface to
a Si--H surface on a silicon wafer through the treatment of the
wafer with a 2.5% aqueous solution of HF. A Si--H surface can also
be formed by other processes, for example by decomposition of
silanes as described in U.S. Pat. No. 6,444,326. A Si--H surface
can also be formed through reacting surface silanol moieties with
reagents such as trihydroxyhydridosilane via sol-gel type methods
(see e.g., U.S. Pat. Nos. 5,017,540, and 5,326,738) or through
dry-etch processes with plasmas of sulfur hexafluoride or Freon 23.
A Germanium hydride (Ge--H) surface can undergo the same types of
the reactions of Si--H to create covalently bonded redox active
reagents. Suitable reactions to covalently bond to the Si or Ge
surface are described, for example, in J. M. Buriak, Chem. Review
2002, 102 (5), 1271. The hydrides of other semiconductor substrates
can also be prepared and used to covalently bond redox active
moieties. Suitable semiconductor hydrides include for example,
hydrides of silicon germanium (M. S. Carroll, et al., J.
Electrochem Soc. 2000, 147 (12), 4652), gallium arsenide (P. E.
Gee, et al., J. Vacuum Sci. Tech. A: Vacuum Surf. Film 1992, 10
(4), 892), gallium nitride, diamond film (S. Yamashita, et al.,
U.S. Pat. No. 5,786,604 (1998)), and indium phosphide (Y. Sun, et
al., J. Appl. Phys. 2005, 97, 124902).
[0258] The semiconductor-hydride surface, e.g., Si--H surface, can
be reacted with a variety of functional groups to create covalent
bonds and thereby attach a redox active moiety to the semiconductor
surface. The Si--H surface, Ge--H surface or other Semiconductor-H
surface can participate in hydrosilylation reactions, involving the
addition of, for example, the Si--H across an unsaturated site to
form a Si--C, Si--O, or Si--N bond to the surface. Functional
groups which can be used in these reactions including
hydrosilylation include alkenes, alkynes, imines, carbonyls and
oximes. These reactions, including hydrosilylation can be carried
out thermally, photochemically, with a metal catalyst or with a
radical initiator (see Buriak, Chem. Commun., 1999, 1051-1060). The
Si--H surface or other semiconductor-H surface can also be reacted
with alkyl or aryl carbanions through, for example, Grignard, or
lithium reagents. In some embodiments, a Si--H surface or other
semiconductor-H surface can react with azido, diazo, and diazonium
groups. Suitable diazonium reactions are described, for example, in
Stewart et al., J. Am. Chem. Soc. 126, 2004, 370-378.
[0259] FIG. 5 illustrates the reaction of a surface Si--H with an
aldehyde functionality attached to a ferrocene redox active moiety
to create a covalently bound ferrocene through a Si--O bond. FIG. 5
also illustrates the reaction of a surface Si--H with a vinyl
functional group attached to a ferrocene redox active moiety to
create a covalently bound ferrocene through a Si--C bond. FIG. 6
illustrates the reaction of a surface Si--H with an aldehyde
functionality attached to an anthracene redox active moiety to
create a covalently bound anthracene through a Si--O bond. FIG. 6
also illustrates the reaction of a surface Si--H with a vinyl
functional group attached to an anthracene redox active moiety to
create a covalently bound anthracene through a Si--C bond. FIG. 7
illustrates the reaction of both a ferrocene redox active moiety
and an anthracene redox active moiety through vinyl functionality
to produce a silicon surface with a covalently attached redox group
that is sensitive to hydrogen ion (anthracene) and a covalently
attached redox group that is insensitive to hydrogen ion
(ferrocene). In some embodiments, a carbonyl group such as an
aldehyde group is substituted for the vinyl functionality for
providing attachment to the surface.
[0260] The redox active moieties can alternatively be attached
covalently to the surface by direct reactions with a semiconductor,
e.g., silicon surface from which all functionality has been
removed, typically by high temperature and vacuum. The pure silicon
surface, for example, can react directly, for example with alkenes
and alkynes to form Si--C covalent attachment. (see Bateman, et.
al., Angew. Chem. Int. Ed. 1998, 37(19), 2683-2685). Diazonium
species can also be used to functionalize the surface either
thermally or electrochemically. In some embodiments, ultrahigh
vacuum techniques can be used to prepare the functionalized
surfaces of the invention, for example by [2+2] Reactions of
alkynes and alkenes or Diels-Alder ([4+2]) reactions of dienes with
reconstructed Si surfaces.
[0261] Capping of semiconductor oxide surfaces such as silica and
glass surfaces with alkyl-, alkoxy- and chloro-silanes may also be
used to functionalize the semiconductor surface.
[0262] The semiconductor surface can contain oxide functionality
including hydroxy functionality (native oxide). In some
embodiments, the semiconductor electrodes of the invention are
modified by covalent attachment to this oxide functionality. For
example, the hydroxy groups of silicon or other semiconductor
elements can be coupled to surface bound groups using the many
reactions known in organic chemistry for carbon bound hydroxy
groups, including for example, the formation of esters and ethers.
One derivatization method involves the use of a carbodiimide for
coupling to the surface. Exemplary carbodiimides include, for
example, dicyclohexylcarbodiimide (DCC), or
(1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride)
(EDC).
[0263] The redox active moieties can also be attached covalently to
the surface by reactions with the native oxide, --O or --OH on the
semiconductor surface. Many methods are known for carrying such
reactions to form covalent bonds through various functional groups
(see Maoz et al., J. Colloid. Interface Sci., 1984, 100, 465-496).
In some embodiments an indirect approach can be employed in which
an alkoxysilane comprising other reactive functionality is reacted
with the --O or --OH groups on the semiconductor surface to
covalently attach the alkoxysilane to the semiconductor surface.
The other reactive functionality on the alkoxysilane can then be
used to covalently attach a redox active moiety to the surface. In
these embodiments, the alkoxysilane can become a linker or portion
of a linker. The other reactive functionality can be any reactive
functionality that can be used to attach a redox active moiety. The
functionality can be, for example, an olefin, an acetylene, an
amine, a mercaptan, or an epoxy group. The reaction used to couple
the alkoxysilane to the redox active moiety can be, for example a,
Diels-Alder reaction, Michael addition, click chemistry, or epoxy
chemistry. Semiconducting polymers comprising redox active moieties
can be polymerized onto a surface, graft polymerized onto a
surface, photo-polymerized, or pre-formed and cast onto a
surface.
[0264] In some embodiments, the reactions described above for
covalently attaching functional groups to the semiconductor surface
are used to attach a linker group or portion of a linker group
having a chemical functionality that can be used to covalently bind
the redox active moiety to the surface in a subsequent step.
[0265] Reactions that can be used to covalently attach a redox
active moiety to the semiconductor surface include hydrosilylation,
free radical reactions, carbodiimide coupling, Diels-Alder
reactions, Michael addition, epoxy reactions, or click chemistry
(see, e.g., Evans et al. Australian Journal of Chemistry 60 (6):
384-395 (2007).
[0266] Organic semiconductor substrates of the invention can in
some embodiments be modified to attach redox active species after
the formation of the organic semiconductor. For example, an organic
semiconductor can be cast into a film. In some embodiments the
organic semiconductive polymers, for example, polyaniline,
polypyrrole, and polythiophene are prepared by chemical or
electrochemical oxidation. The semiconductive polymer can be
subsequently modified using, for example, the chemistry described
for the attachment of redox active agents to the inorganic
semiconducting substrates. In some embodiments, the semiconductive
polymers can comprise functional groups that can be used to attach
redox active species to the semiconductive polymer. For example,
the sulfonic acid groups in poly(anilinesulfonic acid) can be used
to immobilize aminoferrocene, 1,1'-diaminoferrocene, and
aminoanthracene through sulfamide formation. In some embodiments, a
pyrrole, thiophene or aniline monomer can have a functional group
such as an N-hydroxy succinimide ester, a carboxylic acid, or a
reactive vinyl or allyl group. The monomers can then be
polymerized, for example electropolymerized, and the redox active
moiety can be attached to the functional group on the polymerized
monomer.
[0267] In some embodiments, the redox active moiety can be
incorporated into the semiconductive polymer itself; for example as
part of the backbone of the polymer. For example, redox active
groups within poly(ferrocenyl vinylene phenylene vinylene),
poly(fluorine), and poly(carbazole) that comprise redox active
moieties in their main chains, which can be used for analyte, e.g.,
pH sensing.
[0268] In some cases the surface of the semiconductor is modified
by coating the surface with conductive compounds such as metals or
metal oxides. In some embodiments, the semiconductor is coated with
gold, silver, palladium, copper, platinum or other metals. The
metals can be coated from solution, for example by
electrodeposition, or can be coated onto the surface with vacuum
techniques such as plasma deposition, or metal vaporization. The
semiconductor surface can be coated with conductive or
semiconductive metal oxide compounds such as indium-tin oxide. When
these materials are coated onto the semiconductor electrode
surface, the redox-active agents are attached to the semiconductor
electrode by attachment to the layer on the semiconductor
electrode.
[0269] Where a linker group is used, the linker can be small, for
example one to 3 atoms, or can be longer, e.g., 20 to 100 atoms.
The linker can also be any size between the small or longer linker.
In some embodiments, the linker is relatively short allowing for
the redox active moiety to be close to the surface, which can be
beneficial for electron transfer. In some embodiments the linker is
provided such that the redox active agent is held 1, 2, 3, 4, 5, 6,
or 7 atoms from the surface of the semiconductor. Where a short
linker is used, the redox-active moiety is held close to the
surface. Where a longer group is used, the redox active moiety may
be able to move away from the surface, for example further out into
the solution. Linker groups can comprise hydrophilic, hydrophobic
groups, or mixtures thereof. Linker groups can comprise, for
example, hydrocarbons, alkenes, alkynes, esters, ethers, amides,
amines, carbonyls, thiols, olefins, silicones, or other organic,
inorganic or organometallic groups. The linker groups can be formed
by polymerization or oligomerization reactions such as free radical
or anionic polymerization. The linker group can comprise, for
example, ethylene oxide, propylene oxide, or acrylamide repeat
units. Linkers can have ring structures including aromatic rings.
The variation in the linker structure can be used to vary the
mobility of the redox-active moiety in the solution. If the linker
is too long and densely packed the redox active moiety can be far
enough away from the surface such that the electron transfer to the
electrode surface may be compromised. In these cases, having a
linker that has electrical conductivity can be useful.
[0270] A redox-active moiety may be highly substituted, and can
still act as a redox-active moiety. Thus, for example, the
redox-active moiety ferrocene includes substituted ferrocenes,
ferrocene polymers, and ferrocene covalently attached to the
surface via linker molecules.
[0271] In some embodiments, the redox-active moiety can be
incorporated into a polymer, and the polymer comprising the redox
active moiety can be immobilized onto the semiconductor surface.
The immobilization of the polymer can be either chemical or
physical. The immobilization of the polymer can be through covalent
bonds, or through adsorption of the polymer to the semiconductor
surface.
[0272] The redox active moieties can be incorporated into any type
of polymer that can be immobilized onto the surface of the
semiconductor surface. Types of polymers that the redox active
moieties can be incorporated into include biopolymers such as RNA,
DNA or proteins, conductive polymers, fluoropolymers, polyterpenes,
inorganic polymers, phenolic resins polyanhydrides, polyesters,
polyolefins, polysiloxanes, polyamides, polyimides, polyethers,
polyketones, polysulfones, and vinyl polymers.
[0273] The polymer comprising the redox active moieties can, in
some cases, be produced at the semiconductor surface. For example
monomers or oligomers comprising the redox active moieties can be
polymerized in the region of the surface to product the polymer
near the surface. In some cases, the polymerization can be
initiated at the semiconductor surface, resulting in polymer
covalently bound to the surface. The polymerization can be
initiated at the surface can be initiated, for example by a free
radical reaction initiated by a diazo group attached to the
semiconductor surface. In other cases, the polymerization can be
initiated in solution, for example near the surface, such that the
nascent polymer is immobilized onto the surface as it is formed.
Methods for determining the appropriate solvent conditions are
known. For example by establishing that the monomer and/or oligomer
are soluble, while the polymer is insoluble, allowing for surface
deposition to occur. The semiconductor surface can comprise
polymerizable functional groups that are capable of copolymerizing
with the monomers or oligomers comprising the redox active moieties
resulting in covalently binding the redox active polymer onto the
semiconductor surface.
[0274] In some embodiments, a polymer comprising the redox active
moieties can be electropolymerized at the semiconductor surface.
For example, monomers comprising the redox active moieties are
added to a solution, and current is provided through the
semiconductor surface causing electropolymerization of the
monomers. In some embodiments, the electropolymerization can result
in the covalent attachment of the electropolymerized polymer to the
semiconductor surface. In other embodiments, the
electropolymerization can result in polymerization, for example, at
the semiconductor-solution interface, and the polymer that is
formed can deposit onto the semiconductor surface, resulting
immobilization of the polymer by physisorption to the surface. The
polymers can be polymerized onto a surface, graft polymerized onto
a surface, photo-polymerized, or pre-formed and cast onto a
surface.
[0275] The polymers can be chemically or electrochemically
deposited on individual microelectrodes, polymerized, as respond to
a signal in a reversible manner, in a way that can be
electrochemically detected. Other such materials are described by
R. W. Murray in Electroanalytical Chemistry, vol. 13, edited by A.
J. Bard (Marcel Dekker, NY 1984), the teachings of which are
specifically incorporated herein.
[0276] In some embodiments, the polymer is formed away from the
semiconductor surface, and subsequently immobilized thereto. The
polymer can be immobilized onto the surface by a variety of methods
including, adsorption from solution, coating including spin coating
and dip coating, spraying, printing, electropainting, or
electrodeposition.
[0277] In some embodiments the semiconductor electrode is formed by
contacting an H-terminated semiconductor surface with the one or
more redox-active moieties wherein at least one redox active moiety
is sensitive to the presence of an analyte, wherein each
redox-active moiety comprises a functional group that will react
with the H-terminated semiconductor surface to form a covalently
bond, thereby forming a derivatized semiconductor surface. In some
embodiments, the surface comprises at least two redox active
moieties and one of the redox active moieties is insensitive to the
presence of the analyte.
[0278] Methods for forming semidonductor electrochemical sensors
can be applied to forming other solid state electrochemical
sensors, such as carbon sensors. It will be understood that in the
case of other solid state materials, the doping configuration can
be changed to effect desirable sensor properties, such as
sensitivities and reliability.
Uses of the Compositions and Devices
[0279] Another aspect of the invention provides a method for
determining the concentration of an analyte. In an embodiment, the
method comprises bringing an electrode of a sensor in contact with
the analyte, the electrode comprising a solid state (e.g.,
semiconductor) substrate with surface having immobilized thereon an
analyte-sensitive redox-active moiety. The analyte-sensitive
redox-active moiety exhibits an oxidation potential and/or
reduction potential that is sensitive to the analyte. Next, a
plurality of potentials are applied to the electrode. The current
through the electrode is measured at the plurality of potential to
determine a reduction and/or oxidation potential of the
analyte-sensitive redox-active moiety, thereby determining the
concentration of the analyte.
[0280] In some embodiments, the method comprises determining the
concentration of an analyte by (a) placing an electrode in contact
with said analyte, said electrode comprising a solid state (e.g.,
semiconductor) substrate with surface having immobilized thereon an
analyte-sensitive redox-active moiety, said analyte-sensitive
redox-active moiety exhibiting an oxidation potential and/or
reduction potential that is sensitive to the concentration of the
analyte; (b) applying a plurality of potentials to the electrode;
and (c) measuring the current through the electrode at the
plurality of potentials, and determining a reduction and/or
oxidation potential of the analyte-sensitive redox-active moiety,
thereby determining the concentration of the analyte.
[0281] The method of determining the concentration of the analyte
can be used to measure pH by utilizing redox active moieties that
are sensitive to hydrogen ion as described above.
[0282] The measurement of current at a plurality of potentials
allows for carrying out voltammetry for determining the oxidation
and or reduction potential of the redox active moiety or moieties
immobilized on the surface. The voltammetry used in the method can
be, for example cyclic voltammetry, pulse voltammetry, normal pulse
voltammetry, square wave voltammetry, differential pulse
voltammetry linear voltammetry, or square wave voltammetry. The
source for supplying a plurality of potentials can be a
potentiostat, for example, a potentiostat capable of applying
square waves for square wave voltammetry.
[0283] The frequency of the measurement can affect the quality of
the data. In some embodiments, square wave voltammograms are
currently at a step height of 2 mV, amplitude of 25 mV and a
frequency of 10 Hz. In some cases, it is advantageous to increase
the frequency. For example, increasing the frequency to 500 Hz can
result in a faster scan rate. We have observed that in some cases,
a higher frequency results in a higher level of observed current.
In some cases, the peak current can shift, for example to more
negative potentials upon increasing the operating frequency. In
some cases changing the potential at higher frequencies can result
in more noise in the square wave voltammograms. In some cases it is
advantageous to shield the electrode from the light because in some
cases, light can contribute to the background noise.
[0284] In some embodiments, the electrode further comprises an
analyte-insensitive redox-active moiety having a reduction and/or
oxidation potential that is substantially insensitive to the
analyte, and the method further comprising determining the
oxidation and/or reduction potential of the analyte-insensitive
redox-active moiety, and determining the concentration of the
analyte from the difference in the oxidation and/or reduction
potentials of the analyte-sensitive and analyte-insensitive
moieties. The redox-active moiety having a reduction and/or
oxidation potential that is substantially insensitive to the
analyte can be on the same electrode as the redox-active moiety
exhibiting an oxidation potential and/or reduction potential that
is sensitive to the analyte, or a different electrode (e.g.,
separate and electrically isolated electrodes).
[0285] Generally, the analyte concentration is determined by using
voltammetry to identify the position of current peaks, which
current peaks indicate the reduction or oxidation potential of a
redox active moiety. In some embodiments, the position of the
reduction and/or oxidation potential of the analyte sensitive redox
active moiety is used to determine the concentration of the
analyte. For example, the position of the current peak with respect
to the potential at a reference electrode can be used. This method
can be used, for example, where no analyte insensitive redox active
moiety is employed.
[0286] Where an analyte insensitive redox active moiety is used,
detection is generally accomplished by measuring the potential
difference, delta E, associated with current peaks for oxidation
(or reduction) of the immobilized redox active moieties, where the
magnitude of delta E can be related to the concentration of
analyte, e.g., hydrogen ion (H+) in solution. That is, in many
embodiments, delta E represents the potential difference between
the reduction and/or oxidation potential between a redox active
analyte sensitive moiety and a redox active analyte insensitive
redox active moiety. The analyte insensitive redox active moiety
which has an electrochemical response that is insensitive to
variations in the medium serves as the reference. Current peaks for
oxidation or reduction of the reference and indicator can be
determined from a voltammograms using a counter electrode, and
without the need for a reference electrode.
[0287] In some embodiments the measured current through the
electrode at the plurality of potentials is used to determine the
concentration of the analyte. The determination of the
concentration using the measured current (e.g., current peaks) can
be accomplished by using a computation system that communicates
with the device for measuring current. The computation system can
apply algorithms for calculating reduction or oxidation potential
from the measured current at a plurality of potentials from the
voltammetry measurements. The computing systems can be part of the
sensing system, in some cases allowing the sensing system to be
self contained. The computing system can utilize its memory for
storing raw or processed data from the sensors. The method can
further comprise communication between the computing system and the
sensor via transmission device that will wirelessly or by wire
transmit processed data to an external device.
[0288] Carrying out the method typically requires the use of at
least one other electrode (the counter electrode). The counter
electrode is used to complete the electrochemical circuit to make
the measurements described herein. The counter electrode is
generally made of a material that is chemically inert to the medium
so that its potential does not change significantly during the
course of measurement. Suitable materials in many applications
include platinum, gold, stainless steel, and carbon. In some cases,
the counter electrode can be incorporated into the chip that also
comprises the semiconductor sensor electrode.
[0289] A reference electrode is optional and is used as an
additional electrode in some embodiments of the method of measuring
analyte concentration. In the case of a three-electrode system, the
counter electrode generally completes the circuit, allowing current
to flow through the cell, while the reference electrode maintains a
constant interfacial potential difference regardless of the
current. In the case where the system comprises an analyte
sensitive redox active moiety and an analyte insensitive redox
active moiety, the analyte insensitive redox active moiety can act
as a reference, allowing the potential difference to be used to
determine analyte concentration. Even when an analyte insensitive
moiety is also used, in some embodiments, a reference electrode
will still be used. In some embodiments, pseudo-reference
electrodes can also be utilized. Reference electrodes which can be
employed are described above.
[0290] In some embodiments, the sample is a liquid sample, and the
electrodes are each in contact with the liquid. In some cases, the
sample will not be a liquid, but may be a solid, generally
comprising a solid electrolyte, or a gas.
[0291] In some embodiments, the method involves the in-line sensing
of a process. An in-line sensor can be a sensor that is used in an
on-going process. In some embodiments the method comprise the use
of a sensor is in a vessel, in other embodiments the sensor is in a
conduit or pipe through which a process fluid flows. In some
embodiments, the method comprises using currents measured at a
plurality of potentials by voltammetry to determine analyte
concentration, and the determined analyte concentration is used to
control a process parameter. The systems of the present invention
are valuable in in-line sensing in that they can be made to be
robust, to resist fouling, and are able to measure analyte
concentration for long periods of time in media that changes its
properties, as in a process such as a chemical reaction,
biochemical reaction, or fermentation.
[0292] In some cases, under remote monitoring the sensor can be
programmed to automatically take readings. The automatic readings
can be programmed to occur on a periodic basis, to occur upon the
happening of some event, or to occur when the sensor is prompted.
The periodic events can be separated on the order of seconds to on
the order of months. The happening of some event could be, for
instance at the point when the measured solution reaches a certain
volume level, or at given points in the steps of a manufacturing
process (e.g., at the beginning of or end of a step, or upon the
addition of a reagent to a vessel).
[0293] Remote monitoring generally includes communication from the
remote sensing unit, and/or communication to the remote sensing
unit. The communication to and from the remote sensing unit can be
done with transmission lines, and/or wirelessly. Any type of signal
including, for example, digital, analog, wideband, narrowband, or
combinations thereof; can be used.
[0294] Another aspect of the invention provides voltammetric
monitoring of pH with a semiconductor electrode as part of process
control in processes. In an embodiment, a voltammetric pH
measurement is made in an industrial process stream, and the pH
value from that measurement is used to as input to a decision on
the adjustment a process parameter. In an embodiment, the pH value
from the voltammetric pH measurement with a semiconductor electrode
is used to decide whether or not to add one or more components to
the process, and/or to decide how much of the component to add. In
some embodiments, the pH value is used to control the pH in a part
of the process, for example, as input into the decision on the
addition of either acidic or basic components. In some embodiments,
the pH value is used to determine whether a process has reached a
certain stage, for instance, whether a reaction is at completion.
In some embodiments, the pH value is used to determine the addition
of nutrients or other components to a reaction containing an
organism to maintain the health and productivity of the
organism.
[0295] The process control step can be automated such that a given
pH measurement value from the sensor results in the change of a
process parameter without the intervention of a person. In other
embodiments, the pH measurement is viewed by a person who uses the
information to make the decision the change of a process
parameter.
[0296] The process control step can be controlled by a voltammetric
pH system with a semiconductor electrode that has a sensor, a
voltage source, a current measuring detector, and a computer for
determining the pH from the current measurements. The voltammetric
pH system can be in communication either with a process control
system, or with an operator, by analog or digital means, either
with a wire, wireless connections, fibers or combinations
thereof.
[0297] Another aspect of the invention is a method of voltammetric
pH sensing with a semiconductor electrode wherein the pH sensor
requires little calibration. In some embodiments, the pH sensor is
substantially free of the need for calibration or re-calibration.
Substantially accurate measurements may be made without
re-calibration.
[0298] The use of the voltammetric pH sensing with a semiconductor
electrode has a number of advantages. For example, the sensors of
the present invention generally comprise solid-state sensors. The
sensors of the present invention have a built-in internal standard
such that calibration is not required. The sensors of the present
invention can be constructed to be physically robust, such that
they are not prone to breakage. The sensors of the present
invention can be made to be relatively insensitive to fouling. The
sensors of the invention can be constructed to be resistant to
chemical sterilization such as exposure to ethylene oxide, UV
stabilization, gamma irradiation, electron beam irradiation, and
temperature treatment. The sensors of the invention can be
constructed to be resistant to high humidity and high temperature
treatment under pressure such as experienced in an autoclave.
[0299] The voltammetric pH sensing methods with a semiconductor
electrodes comprise reactions carried out in stainless steel
reactors, glass reactors (e.g., for product development), and
disposable reactors (e.g., plastic reagent bags), for example
reactors described by manufacturers such as Wave Biotech, Hyclone,
Xcellerex, and Stedim.
[0300] Another aspect of the invention provides methods for
voltammetric pH sensing with a semiconductor electrode for
processing including chromatography and tangential flow
ultrafiltration.
[0301] Another aspect of the invention is as a sensor in a remote
monitoring system such as a drug (pharmaceutical agent) delivery
system. Such a system is described, for example, in U.S. Patent
Application 2003/0153900, which is entirely incorporated herein by
reference. The analyte monitoring system or monitoring and drug
(pharmaceutical agent) delivery system can be partitioned into a
disposable module, a reusable module and a personal digital
assistant (PDA) module. A PDA is typically a portable, e.g.,
handheld device that has computing and networking capability, and a
user interface, with output, e.g., a display, and input, e.g.,
stylus, keyboard, and/or touchscreen capability. This configuration
optimally distributes functionality among these three
configurations to achieve certain advantages. However the invention
is not limited to this configuration. For example, a one-unit
disposable device including all electronics, microneedles,
chemistry, sensors, mechanics and user interface may be
alternatively employed. Or, more relevantly, the design of the
invention allows for any distribution of components between one or
more system modules. For example, components may be partitioned
among one or more system modules based on the overall system cost,
user safety and/or performance concerns.
[0302] The disposable module contains those components that once
used must be discarded to maintain safety and accuracy. This module
preferably includes any structural or mechanical elements to
maintain integrity, sterility and/or an electromechanical interface
to any reusable components. Therefore this system module can
include, for example: microneedles, a microfluidic assembly,
membrane, reagent chemistry and associate housing materials. The
portion of a sensor which is in contact with a biological fluid,
for example, may be part of the disposable module. This module can
also include retaining mechanisms for establishing and maintaining
intimate contact with the body thereby providing mechanical
retention of the analyte monitoring/drug (pharmaceutical agent)
delivery system.
[0303] The reusable module generally contains those components that
control, automate motion, measure the analyte concentration, alarm
the user, transmit data to the PDA module. This module can also
include retaining mechanisms. Generally, this module includes: a
microprocessor with associated circuitry (e.g., memory, supporting
electronics and the like), sensing circuitry, including, for
example, a voltage supply and current measuring device, drive
mechanisms such as motors or the like, a power supply (e.g.,
battery) and an interface operable to communicate with a portable
computing device or PDA. The interface can be RF, magnetic or
inductive, optical or the like. The reusable module can also have
an audible or vibration alarm to notify the user that user action
intervention is required.
[0304] The PDA module generally includes a separate user interface
via a portable computing device such as a personal digital
assistant (PDA), handheld computer or the like for controlling
and/or interacting with the device. A typical portable computing
device includes a processor, memory, associated circuitry, a
display (e.g., monochrome or color LCD) and an input device such as
a key pad, touch screen (e.g., integrated with the display) or the
like and an operating system. The display can show the value of the
analyte to be measured, could provide the user with instructions on
how to respond to the measured level of analyte, or may tell the
user what automatic actions have been taken in response to a
measured level of analyte.
[0305] Today, portable computing devices with improved operating
system software and user interfaces are readily available. These
devices provide the potential for rich and extended functionality.
For example a typical PDA includes a relatively large viewing
screen and can also include wireless communications mechanisms, a
sophisticated operating system and a variety of business and
personal software (calendars, scheduling, etc.). The invention
preferably includes the use of a PDA to provide the proprietary
software (programs) for autonomous operation with an improved user
interface.
[0306] For example, the PDA module can provide the user with
software that facilitates informed decisions to help a patient user
more optimally adjust either drug levels or behaviors to more
optimal levels. The PDA configuration provides a user interface and
preferably allows users the ability to program and or control
testing. The user can view individual analyte measurements and
graphically display analyte level trends by the day, week or custom
time period. The PDA can be used to display any and all of the
measurements recorded by the system. Using the proper software, the
user can be provided with recommendations for drug regiment
modification. In some cases, the user can program the times when
their analyte tests are to be taken. Preferably, the user can also
set the upper and lower limits for alerts.
[0307] The system can be programmed such that whenever a user makes
changes and with verification from the user, the information can be
wirelessly downloaded to the system. During the day the user may
not need to use the PDA unless alerted by the system to check for
an analyte reading. The user can initiate a test from the PDA if
wanting to make an immediate measurement. Once the user selects
this command, verifies it, and transmits it to the reusable module,
a confirmation is made back to the PDA.
[0308] Another aspect of the invention comprises drug dispenser
capsule comprising a voltammetric sensor. In some embodiments, the
drug dispenser capsule comprises a semiconductor based voltammetric
sensor as described herein. The drug dispenser capsule of the
present invention internally senses a biologic condition by the
detection of the presence or amount of an analyte, and internally
dispenses drugs within the digestive tract of a body (e.g., a human
body or animal body) based upon the sensed level of analyte. The
capsule is inert and is therefore swallowable and passable through
the digestive tract without being consumed. By sensing the level of
one or more analytes, the swallowable drug dispenser capsule senses
information the digestive tract or senses conditions within the
digestive tract that are indicative of conditions in other organs
(e.g., skin). In addition to the voltammetric analyte sensor, the
capsule contains one or more other sensors (e.g., chemical,
electrical, etc.) so that more types of biologic data can be
tracked through the digestive system. In response to that sensed
information, the capsule dispenses a bioactive substance within the
digestive tract without the need to transmit or receive signals
from a remote transmitter/receiver, and without active human or
computer management. Drug dispenser capsules are described, for
example, in U.S. Pat. No. 6,929,636, which is entirely incorporated
herein by reference.
[0309] The swallowable drug dispensing capsule comprising a
voltammetric sensor can include, for example, sensors, a
controller, memory, optional programmable logic, a power supply, a
microactuator, a drug storage module, and communication interface
having at least one of the following types of communication
modules: radiofrequency; ultrasonic; and/or infrared. In one
preferred embodiment, at least memory, and preferably also
controller and/or programmable logic are embodied on a
semiconductor-based, e.g., silicon-based, module in one or more
semiconductor chips.
[0310] In some embodiments, the swallowable drug dispensing capsule
has multiple sensors that are arranged an outer surface of capsule
in a desired predetermined orientation that is expected to expose
each sensor to a targeted bodily condition or landmark within the
human body. Each sensor can comprise a single type of sensor such
as an image detector or a different type of sensor (e.g., chemical,
electrical, temperature, etc.). Chemical detectors detect the
presence of many analytes, such as pH, or other analytes.
[0311] The swallowable drug dispensing capsule of the invention can
have a controller that regulates communication between sensors and
memory, communication between memory and any remote controllers
outside of the human body, and communication with programmable
logic component(s). Finally, controller can operably control both
communication interface and a microactuator. The controller
typically is a logic controller and includes a microprocessor. The
controller may also comprise one or more logical devices (e.g., a
logic gate) capable of performing a sequence of logical
operations.
[0312] The swallowable drug dispensing capsule generally has a
memory or storage device that is typically an ultra-high capacity
storage device, and which is often based on a semiconductor chip,
e.g., a silicon chip.
[0313] The swallowable drug dispensing capsule generally has a drug
storage module and a microactuator. The drug storage module
represents a container for holding a drug or bioactive substance
that may be released, for example, into the digestive tract.
Accordingly, the drug storage module also includes one or more
selectively activated dispensing ports that open in an outer
surface of capsule. The microactuator can have a chemically
activated or electromechanically activated mechanism for causing
the drug storage module to release its contents. The swallowable
drug dispensing capsule has a suitable power supply, such as a
lithium-ion battery, which is relatively non-toxic. Alternatively,
other power supplies that are suitable for in vivo environments can
be used.
[0314] The swallowable drug dispensing capsule generally has a
communication interface that includes any suitable wireless
transmission technology (e.g., ultrasonic, radiofrequency, etc.)
that readily permits communication to and from the capsule while
the capsule is in digestive tract and the remote
transmitter/receiver which is located remotely outside of the body.
However, a wireless port is preferably used for communicating with
capsule after capsule is captured from the body. Likewise, a
wireless port may be used for programming the controller, memory,
and/or logic component prior to insertion of capsule within the
body to determine the manner that the sensors will operate and
communicate with the memory, as well as the manner that
microactuator will operate and communicate with memory via
controller.
[0315] In use, the sensors, including the voltammetric sensor of
the capsule sense analyte concentrations and biologic data within
the digestive tract and the sensed data is passed through the
controller for storage in memory and/or comparison with a stored
data profile in memory and/or logic. After the predetermined
criteria are met, controller activates microactuator to dispense
the drug from drug storage module into digestive tract. The sensed
data optionally is stored in memory and retrieved via the
communication interface after capture of capsule upon exiting the
digestive tract. Finally, a wireless communication system
optionally can be used in addition to, or as an alternative to,
controller and memory to facilitate evaluating and storing sensed
data and to dispense drugs upon selective activation at the
appropriate time.
[0316] The semiconductor electrode voltammetric pH sensors of the
present invention can be used in manufacturing operations such as
the manufacture of coatings, cleaners and sealers that enhance
paint and finish bonding, metal passivation to protect substrates
during shipment and storage, paint spray booth treatments that
enhance quality and efficiency, and air scrubbers that limit
pollutant emissions. In these applications, the reliable
measurement of pH can be an integral part of the process.
[0317] In some embodiments, sensors can be used as embeddable
corrosion measuring instruments that are capable of providing
information related to corrosion rate, corrosion potential,
conductivity and chloride concentration, and/or pH levels of steel
rebar reinforced structures. The devices can be used to monitor the
integrity of the steel. The devices and systems of the present
invention do not require a direct electrical connection to the
reinforcement steel within the structure, using the structural
steel as one of the referencing materials. Since the disclosed
instruments do not require proximity to the steel within the
structure, the instruments can be dispersed at critical locations
within the structure, regardless of steel placement. In some
embodiments, the systems and devices are self-contained,
incorporating all required sensing electrodes and electronics. The
devices can be deployed as described in U.S. Pat. No. 6,690,182,
which is entirely incorporated herein by reference.
[0318] The semiconductor electrode voltammetric pH sensors of the
present invention can be used in winemaking. Measurements of
various properties including pH are taken throughout the process,
including during (1) pressing, (2) primary fermentation, which
often takes between one and two weeks, where yeast converts most of
the sugars in the grape juice into ethanol (alcohol) (3) secondary
fermentation.
[0319] The semiconductor electrode voltammetric pH sensors of the
present invention can be used in brewing. The measurement of pH can
be important at the various stages of brewing, for example, at
mashing, lautering, lauter tun, mash filter, boiling, whirlpool,
wort cooling, fermenting, conditioning, filtering, and secondary
fermentation. The semiconductor electrode voltammetric pH sensors
of the present invention can be particularly important during
fermentation, where the voltammetric pH sensors of the present
invention are advantageous as they require little to no
calibration, and can be made to resist fouling during the
fermentation process.
[0320] The semiconductor electrode voltammetric pH sensors of the
present invention can be used in the production of biofuels,
including the production of biodiesel, ethanol, butanol, and
substitutes for gasoline, diesel, jet fuel, and additives to be
used in any of the forgoing. The production of ethanol includes
both the process of converting the cellulose to sugars, and the
process of converting the sugars to ethanol. Although there are
several key technological differences in how ethanol is produced
from corn or cellulosic feedstock, both paths to ethanol production
typically require a fermentation step that involves the conversion
of glucose and other sugars to ethanol. Currently, baker's yeast,
Saccharomyces cerevisiae, provides the primary microbiological
system used by the corn-based ethanol industry. The methods of the
present invention relate to ethanol production for fuel from
Saccharomyces cerevisiae and other organisms. The control of pH can
be useful in catalytic biofuel production processes, such as for
the production of biofuel.
[0321] The semiconductor electrode voltammetric pH sensors of the
present invention can be used in oil recovery and refining. The
sensors can be incorporated into down-hole devices for measuring
the analytes present in the down-hole environment. The sensors can
be used in other aspects of processing the oil such as in oil
refining.
[0322] The semiconductor electrode voltammetric pH sensors of the
present invention can be used in the production of
biopharmaceuticals, for example, medical drugs produced using
biotechnology. They include, for example, proteins (including
antibodies), nucleic acids (DNA, RNA or antisense oligonucleotides)
used for therapeutic or in vivo diagnostic purposes.
Biopharmaceuticals are produced by means other than direct
extraction from a native (non-engineered) biological source. An
example is recombinant human insulin (rHI, trade name Humulin),
which was developed by Genentech and marketed by Eli Lilly.
[0323] Another aspect of the invention is a semiconductor electrode
voltammetric pH sensor for the production of biopharmaceuticals
including: blood factors (e.g., Factor VIII and Factor IX),
thrombolytic agents (e.g., tissue plasminogen activator), hormones
(e.g., insulin, growth hormone, gonadotrophins), haematopoietic
growth factors (e.g., erythropoietin, colony stimulating factors),
interferons (e.g., interferons-.alpha., -.beta., -.delta.),
interleukin-based products (e.g., interleukin-2), vaccines
(Hepatitis B surface antigen), monoclonal antibodies (e.g.,
infliximab, basiliximab, abciximab, daclizumab, gemtuzumab,
alemtuzumab, rituximab, palivizumab, trastuzumab (herceptin), and
etanercept) and other products such as tumor necrosis factor, and
therapeutic enzymes.
[0324] Another aspect of the invention is a method for forming a
protein comprising carrying out a fermentation reaction that
produces such protein, wherein the pH of the fermentation reaction
is controlled by measuring the pH with a pH sensor comprising a
solid state (e.g., semiconductor) electrode with a surface having
immobilized thereon a redox active moiety that is sensitive to the
presence of hydrogen ion, and using the measured pH to control the
pH of the fermentation reaction. The solid state electrode in some
cases is formed of a semiconductor, such as silicon. In some
embodiments, the control of the pH can be manual, for instance,
where an operator reads the pH from the pH sensor and uses the
measured pH to determine whether or how much to adjust the pH, and
in other embodiments, the control can be automatic, where the pH
measurement is read by instruments that can adjust the pH based on
the value of the measurement received.
[0325] The semiconductor electrode voltammetric pH sensors of the
present invention can be used for biopharmaceuticals produced from
microbial cells (e.g., recombinant E. coli), mammalian cell lines
and plant cell cultures in bioreactors of various
configurations.
[0326] Cell culture requires cells to be grown, often under a
strict set of conditions to maintain the health of the cells and
maximize the production of the culture. Cells are grown and
maintained at an appropriate temperature and gas mixture (for
example, 37.degree. C., 5% CO2) in a cell incubator. Culture
conditions vary widely for each cell type, and variation of
conditions for a particular cell type can result in different
phenotypes being expressed.
[0327] Aside from temperature and gas mixture, the most commonly
varied factor in culture systems is the growth medium. Recipes for
growth media can vary in pH, glucose concentration, growth factors,
and the presence of other nutrient components. The effect of
changes in pH can be dramatic in some cases, and it can be
important to maintain the pH. The devices, systems, and methods of
the invention allow for control of pH within a range of 1, 0.5,
0.02, 0.1, 0.05, 0.02, 0.01 pH units or less to maintain the growth
and health of the cells. The semiconductor electrodes of the
present invention allow for the accurate measurement of pH with
limited fouling, and in some embodiments, no need for
calibration.
[0328] The semiconductor electrode voltammetric pH sensors of the
present invention can be used for cells that are grown completely
in solution, and for cells that are grown on a substrate. Some
cells naturally live without attaching to a surface, such as cells
that exist in the bloodstream. Others require a surface, such as
most cells derived from solid tissues. Cells grown unattached to a
surface are referred to as suspension cultures. Other adherent
cultures cells can be grown on tissue culture plastic, which may be
coated with extracellular matrix components (e.g., collagen or
fibronectin) to increase its adhesion properties and provide other
signals needed for growth.
[0329] Another aspect of the invention is a bioreactor or fermentor
in which the reaction or fermentation occurring therein is
controlled by a semiconductor based voltammetric sensor of the
invention. In an embodiment, the invention comprises a bioreactor
comprising a semiconductor based sensor wherein the semiconductor
sensor comprises a semiconductor surface having immobilized thereon
a redox active moiety that is sensitive to the presence of an
analyte, such as hydrogen ion. FIG. 17 shows an example of a
bioreactor of the invention comprising a probe for measuring pH,
and thereby controlling the pH in the reactor during the reaction.
The probe comprises an electrode having a semiconductor surface
having immobilized thereon a redox active moiety that is sensitive
to hydrogen ion. In some embodiments the probe comprises two
electrodes, each comprising a semiconductor surface, one of the
electrodes having immobilized thereto a redox active moiety that is
sensitive to hydrogen ion, and one of the electrodes having
attached thereto a redox active moiety that is insensitive to
hydrogen ion. In some embodiments the probe further comprises a
counter electrode, and in some embodiments it further comprises a
reference electrode.
[0330] The semiconductor electrode voltammetric pH sensors of the
present invention can be used to assist in the successful
manipulation of cultured cells. As cells generally continue to
divide in culture, they generally grow to fill the available area
or volume. This can generate several issues that the reliable
measurement of pH can assist with, such as: Nutrient depletion in
the growth media; accumulation of apoptotic/necrotic (dead) cells;
cell-to-cell contact stimulating cell cycle arrest, causing cells
to stop dividing known as contact inhibition; cell-to-cell contact
stimulating promiscuous and unwanted cellular differentiation
Sometimes these issues can be identified by monitoring pH, alone or
in combination with other measurements, and can then be controlled
or remediated by adjusting tissue culture conditions that often
rely on sterile techniques. These methods aim to avoid
contamination with bacteria or yeast that will compete with
mammalian cells for nutrients and/or cause cell infection and cell
death. The pH measurements of the present invention are amenable to
being carried out in a biosafety hood or laminar flow cabinet to
exclude contaminating micro-organisms.
[0331] The semiconductor electrode voltammetric pH sensors of the
present invention can be used for pH sensing in plant tissue
culture, bacterial and yeast cell culture, and viral cell
culture.
[0332] Another aspect of the invention is a semiconductor electrode
voltammetric pH sensor for sensing pH in plant tissue culture. The
pH measurements of the present invention can be used at any step of
plant cell culture. Plant tissue culture is typically performed
under aseptic conditions under filtered air. Living plant materials
from the environment are naturally contaminated on their surfaces
(and sometimes interiors) with microorganisms, so surface
sterilization of starting materials (explants) in chemical
solutions (usually alcohol and mercuric chloride) is an important
first step. Explants are then usually placed on the surface of a
solid culture medium, but are sometimes placed directly into a
liquid medium, particularly when cell suspension cultures are
desired. Solid and liquid media are generally composed of inorganic
salts plus a few organic nutrients, vitamins and plant hormones.
Solid media are prepared from liquid media with the addition of a
gelling agent, usually purified agar. The pH measurements of the
present invention can be made in the liquid, in the moist soil, or
in the agar. The composition of the medium, particularly the plant
hormones and the nitrogen source (nitrate versus ammonium salts or
amino acids), and the pH, can have profound effects on the
morphology of the tissues that grow from the initial explant. For
example, an excess of auxin will often result in a proliferation of
roots, while an excess of cytokinin may yield shoots.
[0333] The semiconductor electrode voltammetric pH sensors of the
present invention can be used with any cell line including:
National Cancer Institute's cancer cell lines, zebrafish ZF4 and
AB9 cells, Madin-Darby Canine Kidney MDCK epithelial cell line,
Chinese Hamster Ovary CHO cells, Insect cell line Sf21, MCF-7
(breast cancer), MDA-MB-438 (breast cancer), U87 (glioblastoma),
A172 (glioma), HeLa (cervical cancer), HL60 (promyelocytic
leukemia), A549 (lung cancer), HEK 293 cells (kidney--original HEK
line is contaminated with HeLa), SHSY5Y Human neuroblastoma cells,
cloned from a myeloma, Jurkat cell line, derived from a patient
with T cell leukemia, BCP-1 cells (PEL), Primate cell lines, Vero
(African green monkey Chlorocebus kidney epithelial cell line
initiated 1962), COS-7 (African Green Monkey Kidney Cells), Rat
tumor cell lines, GH3 (pituitary tumor), 9L (glioblastoma), Mouse
cell lines, 3T3 cells, MC3T3 (embryonic calvarial), C3H-10T1/2
(embryonic mesenchymal), NTH-3T3 (embryonic fibroblast),
Invertebrate cell lines, C6/36 Aedes albopictus (Asian tiger
mosquito) larva, Plant cell lines, Tobacco BY-2 cells (kept as cell
suspension culture, they are model system of plant cell).
[0334] Another aspect of the invention is a semiconductor electrode
voltammetric pH sensor for use in water purification. Water
purification is the process of removing contaminants from a raw
water source, the goal is generally to produce water for a specific
purpose with a treatment profile designed to limit the inclusion of
specific materials. Water purification is not only water purified
for human consumption or drinking water. The semiconductor
electrode voltammetric pH sensors of the present invention can also
be used water purified to meet the requirements of medical,
pharmacology, chemical and industrial applications. The
semiconductor electrode voltammetric pH sensors of the present
invention can be used in water purification processes including,
but not limited to ultraviolet light; filtration; water softening;
reverse osmosis, ultrafiltration; molecular stripping;
deionization; and carbon treatment. Water purification may remove
particulate sand, suspended particles of organic material;
parasites, such as giardia; cryptosporidium; bacteria; algae;
virus; fungi, etc; minerals such as calcium, silica, magnesium; and
toxic metals such as lead, copper; and chrome. Some purification
may be elective in its inclusion in the purification process;
examples, smell (hydrogen sulfide remediation), taste (mineral
extraction), and appearance (iron encapsulation).
[0335] Water from any source is applicable to the present
invention. Groundwater (well water) is an economical choice for
drinking water, as it is inherently pre-filtered, by the aquifer
from which it is extracted. Water from an aquifer will have a
limited output and can take thousands of years to recharge. Surface
water; (rivers, lakes, streams) is far more abundant and is the
typical raw water source used to make drinking water, as a water
source it is carefully monitored for the presence of a variety of
contaminants. The methods of the present invention encompass the
voltammetric measurement of pH of these types of water where the pH
value of the measurement can be used to decide on the purity of the
water.
[0336] The semiconductor electrode voltammetric pH sensors of the
present invention can be used with water purification methods
including: pumping and containment, screening, storage,
pre-conditioning, pre-chlorination, and removal of the fine solids,
micro-organisms and some dissolved inorganic and organic
materials.
[0337] Distilled water generally has an average pH of about 7
(neither alkaline nor acidic) and seawater generally has an average
pH of 8.3 (slightly alkaline). If the water is acidic (lower than
7), lime or soda ash can be added to raise the pH. Lime is the more
common of the two additives because it is cheaper, but it also adds
to the resulting water hardness. Neutralizing with soda ash,
however, increases the sodium content of the water. Making the
water slightly alkaline helps ensure that coagulation and
flocculation processes work effectively and also helps to minimize
the risk of corrosion in pipes and pipe fittings. The pH value can
be used to determine whether water is likely to be hard or soft. In
general, water with a low pH (<6.5) is acidic, and tends to be
soft, and corrosive. Therefore, the water could contain metal ions,
such as iron, manganese, copper, lead, and zinc. In some cases this
results in elevated levels of toxic metals. This can cause
premature damage to metal piping, and have associated aesthetic
problems such as a metallic or sour taste, staining of laundry, and
the characteristic "blue-green" staining of sinks and drains. More
importantly, there are health risks associated with these ions or
contaminants. The primary way to treat the problem of low pH water
is with the use of a neutralizer. The neutralizer feeds a basic
solution into the water to prevent the water from reacting with the
household plumbing or contributing to electrolytic corrosion. Water
with a pH>8.5 could indicate that the water is hard. Hard water
does not pose a health risk, but can cause aesthetic problems.
These problems include an alkali taste to the water, formation of a
deposit on dishes, utensils, and laundry basins, difficulty in
getting soaps and detergents to lather, and formation of insoluble
precipitates on clothing.
[0338] Another aspect of the invention is sensing of analyte levels
such at the pH of bodies of water for example for resource control.
The body of water can be, for example, a lake, ocean, stream, or
river. The ability of the invention to be used remotely and to be
used without the need for frequent calibration or any calibration
at all allows the systems, devices, and electrodes to be deployed
remotely in bodies of water to provide information on analytes such
as hydrogen ion, etc. in such remote bodies.
[0339] Another aspect of the invention is a semiconductor electrode
voltammetric pH sensor for the measurement of pH in processes
related to sewage treatment. Sewage treatment can have the same
steps as described above, but may refer to water that has a higher
level of contamination. Raw influent (sewage) can be the liquid and
semi-solid waste from toilets, baths, showers, kitchens, sinks etc.
Household waste that is disposed of via sewers can compose the
sewage. In some areas sewage also includes some liquid waste from
industry and commerce.
Stability Control
[0340] In some cases, traditional electrochemical sensors can
suffer from IV fluctuations during measurement, which can lead to
inaccurate measurements and low measurement times as more
measurements would have to be taken to obtain a time-averaged
measurement. Such fluctuations can lead to instability in square
wave voltammetry (SWV).
[0341] In some cases, exposing a ferrocene functionalized silicon
surface to an alkaline environment can shift the reaction
Fe.sup.2+Fe.sup.3++e.sup.- away from equilibrium. The Fe.sup.3+
(ferricenium) ions can react with the anion in the alkaline
solution and lose the ability to reverse back to Fe.sup.2+
(ferrocenium) ions. This can adversely affect the stability of an
electrochemical sensor having a ferrocene-containing working
electrode.
[0342] In some embodiments, an electrochemical sensor is provided
having one or more working electrodes comprising species that are
active to an analyte of interest, the species disposed over a solid
state (e.g., semiconductor) surface, such as a silicon surface. One
or more of the working electrodes can be covered by a layer of a
polymeric material. In an embodiment, one working electrode
comprises a layer of a redox-active moiety and a layer of polymeric
material over the layer of a redox-active moiety. In another
embodiment, an electrochemical sensor comprises two working
electrodes with each working electrode comprising a layer of a
redox-active moiety and a layer of polymeric material over the
layer of the redox-active moiety. The electrochemical sensor can
further include a counter electrode and a reference electrode (as
described above). In an embodiment, the electrochemical sensor
includes two working electrodes, a first working electrode having a
redox-active moiety that is sensitive to an analyte of interest
(e.g., H+) and a second working electrode having a redox-active
moiety that is insensitive to an analyte of interest. In an
embodiment, the electrode having a redox-active moiety that is
insensitive to an analyte comprises a layer of a polymeric
material. In some embodiments, the layer of the polymeric material
can include a homopolymer of a copolymer. In an embodiment, the
layer of the polymeric material can include a
fluoropolymer-copolymer. In another embodiment, the layer of the
polymeric material can include a sulfonated tetrafluoroethylene
based fluoropolymer-copolymer. In another embodiment, the layer of
the polymeric material can include perfluorosulfonate inonomer or a
derivative of perfluorosulfonate inonomer. In another embodiment,
the layer of the polymeric material can include Nafion, having the
formula C.sub.7HF.sub.13O.sub.5S*C.sub.2F.sub.4, and the following
structure:
##STR00001##
The layer of polymeric material, such as a Nafion membrane (or film
or layer), can advantageously stabilize its response in square wave
voltammetry (SWV). This can aid in preventing peak potentials of
the working electrode from changing with pH, which could otherwise
cause deterioration of sensor performance. In addition, the layer
of polymeric material can protect the semiconductor surface of the
electrochemical sensor against changes in solution pH, which can
stabilize the peak position for this electrode in SWV. In an
embodiment, in cases in which a working electrode comprises
ferrocene, the layer of polymeric material can selectively screen
off anions and only allow the passage (or flow) of cations, which
can minimize the interaction of anions and ferricenium ions,
thereby preserving the integrity of the redox reaction.
[0343] In an embodiment, an electrochemical sensor is provided
having a working electrode comprising a redox active moiety and a
layer of polymeric material over the redox active moiety. The layer
of polymeric material can include Nafion. The layer of polymeric
material can have a thickness between about 1 nanometer ("nm") and
1000 micrometers ("microns"), or between about 100 nm and 500
microns, or between about 200 nm and 250 microns, or between about
500 nm and 125 microns.
[0344] In an embodiment, an electrochemical sensor is provided
having a working electrode having a redox-active moiety disposed
thereon, wherein the redox-active moiety is sensitive or
insensitive to an analyte of interest. A Nafion-containing membrane
(or layer) is disposed over the working electrode. In an example,
an electrochemical sensor is provided having a working electrode
comprising a Nafion-containing membrane over the working electrode,
the working electrode having a layer of one or more redox-active
moieties.
[0345] In an embodiment, an electrochemical sensor is provided
having a working electrode comprising a layer of ferrocene, and a
layer of a polymeric material over the layer of ferrocene. The
layer of polymeric material can include Nafion. The layer of
polymeric material can have a thickness between about 1 nanometer
("nm") and 1000 micrometers ("microns"), or between about 100 nm
and 500 microns, or between about 200 nm and 250 microns, or
between about 500 nm and 125 microns.
[0346] In another embodiment, an electrochemical sensor is provided
having a working electrode comprising a layer of anthracene, and a
layer of a polymeric material over the layer of anthracene. The
layer of polymeric material can include Nafion. The layer of
polymeric material can have a thickness between about 1 nanometer
("nm") and 1000 micrometers ("microns"), or between about 100 nm
and 500 microns, or between about 200 nm and 250 microns, or
between about 500 nm and 125 microns.
[0347] A working electrode can include a first layer of polymeric
material, such as Nafion, and a second layer of polymeric material
over the first layer of polymeric material. The second layer of
polymeric material may be a protective layer, which may protect the
first layer of polymeric material and the working electrode from,
for example, corrosion, or damage due to impact. In an example, a
working electrode includes a Nafion-containing (e.g., a porous
material impregnated with Nafion) over a layer of redox-active
moieties, and a layer of a second polymeric material over the
Nafion-containing layer. In an example, the second layer of
polymeric material includes polyethersulphone (PES).
[0348] In some cases, the second layer of polymeric material can be
a light blocking layer (see below). The light blocking layer may
aid in minimizing, if not eliminating, the interaction of the
working electrode with light during operation.
[0349] In some embodiments, an electrode, such as a working
electrode, can comprise a layer of a composite material over the
electrode. The composite material can include a porous material,
such as porous plastic or porous silica, which has been treated
with a polymeric material, such as Nafion (or other
fluoropolymer-copolymer). The porous material in some cases is
impregnated with Nafion. In some situations, the Nafion-containing
layer selectively filters out negative ions (anions) that may
interfere with the operation of the electrode. In some situations,
the use of a porous material may advantageously provide thickness
uniformity of the Nafion layer over the electrode.
[0350] In some embodiments, the layer of the composite material,
such as a membrane containing porous plastic and Nafion, has a
thickness between about 1 nanometer ("nm") and 1000 micrometers
("microns"), or between about 100 nm and 500 microns, or between
about 200 nm and 250 microns, or between about 500 nm and 125
microns. In some embodiments, a porous material used in the
preparation of the composite plastic membrane (or film, or layer),
such as porous plastic or porous silica, has one or more pores at
pore sizes (diameters) between about 1 micron and 1000 microns, or
between about 10 microns and 500 microns, or between about 50
microns and 100 microns. In some embodiments, the Nafion content in
the pores of the porous material used in the preparation of the
composite material, such as porous plastic or porous silica, is
between about 1% and 99%, or between about 10% and 50%, or between
about 35% and 65%. The porous composite material can be prepared by
impregnation or sol-gel synthesis.
[0351] In another aspect of the invention, an electrode comprises a
layer of a light blocking material that reduces, if not eliminates,
the interaction of light with a surface of the electrode, such as a
working electrode. Use of a light blocking layer on electrodes has
led to the unexpected realization that device performance during
device use in ambient conditions (such as upon exposure to sunlight
or room light) may be improved if incident light on a surface of a
working electrode (and in some cases reference and counter
electrode) is minimized, if not eliminated. The light blocking
layer in some cases is covered with a layer of a protective
material, such as PES. In some embodiments, an electrode having a
light blocking layer comprises a light emitting device (see below)
on a side of the electrode opposite from the light blocking layer.
In other embodiments, the light blocking layer is formed of
polyethersulphone (PES), such as an opaque PES or PES that does not
transmit light. In some cases, the PES is "black" PES. In some
situations, the light blocking layer is a porous material that is
optically opaque or otherwise blocks light from reaching a surface
of an electrode. The porous material permits an analyte to reach a
surface of the electrode while minimizing, if not eliminating, the
interaction of light with the surface of the electrode. Examples of
such light blocking layers include polymeric materials, such dark
porous plastics. In an example, a dark porous plastic includes
PES.
[0352] In some embodiments, an electrode (e.g., working electrode,
reference electrode, counter electrode) comprises a light blocking
layer for minimizing the interaction of the electrode and/or light
sensitive moieties on or over the electrode with light, such as
ambient light. The light blocking layer may be formed of a
polymeric material, such as polyethersulphone (PES) (e.g., black
PES). In some situations, a light blocking layer may advantageously
screen off light without impeding the flow of an analyte (e.g.,
hydrogen ions), or impeding the interaction of the analyte with the
surface of the electrode. In some situations, the light blocking
layer can prevent (or minimize) light from inducing electron
excitation on the semiconductor surface of the working electrode,
which may interfere with the operation of the working
electrode.
[0353] The light blocking layer may include one or more pores for
permitting an analyte from reaching a surface of the electrode,
such as a surface of a working electrode having redox active
moieties. The light blocking layer may have pores at a pore size
(diameter) between about 0.1 micrometers ("microns") and 1 micron,
or 0.2 microns and 0.9 microns, or 0.45 microns and 0.8
microns.
[0354] In some embodiments, the light blocking layer can prevent
between about 50% to about 99.9% of light, or between about 60% and
90% of light, or between about 70% and 80% of light from reaching a
surface of an electrode, such as a semiconductor surface that may
exhibit sensitivity to light. In some cases, the light blocking
layer transmits less than 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%,
3%, 2%, 1%, or lower of light incident on the light blocking layer.
The percentage of light transmitted through the light blocking
layer is proportional to the percentage of light that reaches a
surface of an electrode.
[0355] In another aspect of the invention, electrochemical sensors
are provided having electrodes operatively coupled to light
emitting devices. In some examples, electrodes are formed on, or
are included in, light emitting devices that are configured to
generate light. Light emitting devices in some cases are light
emitting diodes (LED's).
[0356] In some situations, it has been observed that
electrochemical sensors exposed to light sources of variable
wavelength and/or intensity, such as ambient light (e.g., sunlight,
room light) produces unpredictable or erratic signals (e.g.,
noise), which may adversely affect device performance. However, it
has been observed that operatively coupling electrodes (e.g.,
working electrodes, reference electrode and counter electrode) to a
fixed wavelength, fixed intensity light source, such as a light
emitting device (also "light-emitting device" herein), unexpectedly
minimizes, if not eliminates such erratic signals during
electrochemical measurements. In some cases, exposing an
electrochemical sensor to light during use provides for improved
sensor operation when the electrode is exposed to ambient light.
This advantageously precludes the need to operate the sensor in the
dark, for example.
[0357] In some embodiments, during use, an electrode is saturated
with light from a light source. In other embodiments, during use,
an electrode is exposed to light such as electrons and holes
generated in the electrode upon exposure to light are at least
about 1%, or 10%, or 20%, or 30%, or 40%, or 50%, or 60%, or 70%,
or 80%, or 90%, or 95%, or 99% of the maximum concentration of
electrons and holes that may be generated in the electrode upon
exposure to light.
[0358] In some embodiments, a working provided herein includes a
layer of a redox active moiety adjacent a surface of the electrode,
and a light blocking layer over the layer of the redox active
moiety. The light blocking layer in some cases substantially covers
the layer of the redox active moiety.
[0359] In some embodiments, electrochemical sensors are provided in
which one or more electrodes, such as a working electrode, is
formed on a light emitting device, such as a light emitting diode,
or a plurality of electrodes are formed on a light emitting device.
In such a case, light from the light emitting device is incident on
an underside of one or more electrodes, and may in some cases
propagate through at least a portion of the one or more electrodes.
The ohmic contact side of the one or more electrodes may not be
configured to come in contact with a solution having an analyte
(e.g., H.sup.+).
[0360] In other embodiments, electrodes may not be formed on light
emitting devices, but surface of electrodes (e.g., a surface of a
working electrode having a redox active moiety) is in view of a
light emitting device. In such a case, light from the light
emitting device is incident on the surface of the electrode
configured to come in contact with a solution having an
analyte.
[0361] In some embodiments, the working electrode can be
operatively coupled to a light source below or above a surface of
the working electrode. Light from the light source may minimize, if
not eliminate, interactions of light with a surface of the working
electrode. In some embodiments, the light source comprises a lamp,
a light-emitting diode (LED), or other light emitting device that
illuminates a top surface (having redox active moieties) of the
working electrode.
[0362] In other embodiments, the light source illuminates an
underside of the working electrode. The underside may not have
redox active moieties. In such a case, the working electrode may be
sufficiently thin to permit light to propagate through the working
electrode.
[0363] FIG. 29 shows a working electrode having a light-emitting
device, such as a light emitting diode, and an electrode adjacent
to the light-emitting device. In some embodiments, the
light-emitting device is a light emitting diode (LED) having an
active material (or active region) configured to generate light
upon the recombination of electrons and holes. In some embodiments,
the working electrode is formed of a semiconducting material, such
as silicon. The working electrode can be formed over the LED layer
using, for example, spincoating or vapor deposition techniques,
such as chemical vapor deposition or atomic layer deposition. The
surface of the working electrode further comprises a redox-active
moiety immobilized thereon, said redox-active moiety exhibiting an
oxidation potential and/or a reduction potential that is sensitive
or insensitive to the presence of an analyte. In some embodiments,
the redox-active moiety can be immobilized on the semiconducting
layer by reactions such as hydrosilylation, free radical reactions,
carbodiimide coupling, Diels-Alder reactions, Michael addition,
epoxy reactions, or click chemistry.
[0364] In some embodiments, the working electrode comprises a
non-semiconducting material, such as carbon. In some embodiments, a
layer of non-semiconducting material can be combined with the LED
layer by, for example, spraycoating or vapor deposition techniques.
The surface of the working electrode further comprises a
redox-active moiety immobilized thereon, said redox-active moiety
exhibiting an oxidation potential and/or a reduction potential that
is sensitive or insensitive to the presence of an analyte. In some
embodiments, the redox-active moiety can be immobilized on the
non-semiconducting layer, such as activated carbon, by first
oxidatively treating the non-semiconducting layer to create oxygen
groups on the layer (Lemus-Yegres et al., Microporous and
Mesoporous Materials 109 (2008), 305-316). The non-semiconducting
layer can then be subjected to immobilization reactions.
[0365] In some cases, the LED may minimize, if not eliminate,
interactions of light with the surface of the working
electrode.
[0366] In an example, a working electrode includes a light emitting
diode having a p-type Group III-V semiconductor material, an active
layer (e.g., multiple quantum well active layer adjacent to the
p-type Group III-V semiconductor material), and an n-type Group
III-V semiconductor material layer adjacent to the active layer. A
first electrode is formed adjacent to the p-type Group III-V
semiconductor material. A second electrode is formed adjacent to
the n-type Group III-V semiconductor material layer. The second
electrode may be formed over the n-type Group III-V semiconductor
material layer with the aid of a transition layer, such as indium
tin oxide. The electrode may be formed of one or more transition
metals, such as gold and/or silver. A layer of an insulating
material is then formed on the electrode. In some cases, the layer
of the insulating material is formed of a metal oxide or a nitride
(e.g., silicon nitride). Next, a third electrode is formed on the
layer of the insulating material. The third electrode will serve as
the back contact to the working electrode. Next, the working
electrode is formed on the third electrode. Redox active moieties
are then provided on a surface of the working electrode. The
working electrode may be formed of a semiconducting (e.g., silicon,
germanium, gallium nitride) or non-semiconducting (e.g., carbon, a
metal) material.
[0367] In some embodiments, a light emitting device adjacent an
electrode, such as a working electrode, emits light having a
wavelength greater than or equal to about 300 nanometers (nm), 400
nm, 500 nm, 600 nm, 700 nm, or 800 nm. In some situations, the
light emitting device emits light having a wavelength above 750 nm
or 800 nm. In an example, the light emitting device emits near
infrared or infrared light.
[0368] Light may be exposed to sensor surfaces having redox active
moieties, or sensor surfaces opposite from the redox active
moieties (e.g., back surfaces of a sensor). In some cases, the
thickness of a working electrode is selected to provide a desirable
sensor output when exposed to light from the backside. In some
embodiments, a working electrode has a thickness between about 100
nanometers ("nm") and 1 millimeter, or between about 500 nm and 750
micrometers ("microns"), or between about 250 microns and 650
microns.
[0369] In some embodiments, exposure of a working electrode to
light, such as, e.g., a fixed intensity, fixed wavelength light,
provides improved sensor signal to noise. In some cases, the signal
to noise is improved by a factor of 10, 100, 1000, 10,000, 100,000,
or more.
[0370] The wavelength of light emitted by the light emitting
device, in some cases, is adjustable. In some cases, the wavelength
of light emitted by the light emitting device is adjusted based on
the power applied to the light emitting device. In some cases, the
sensor has an ambient light sensor that detects the level of
ambient light, and adjusts the wavelength of light emitted by the
light emitting device. In an example, if the ambient light sensor
detects little to no ambient light, the light emitting device will
not be used. In another example, if the ambient light sensor
detects light above a predetermined threshold, then the light
emitting device may be used.
[0371] Light emitting devices provided herein may be controlled
with the aid of processors and software. The processor may be an
on-board processor, such as mounted on a printed circuit board in
proximity to the sensor and any ambient light sensor. The processor
may aid in regulating the functionality of any of the sensor, the
light emitting device, and the ambient light sensor.
Co-Functionalization of Working Electrodes
[0372] In another aspect of the invention, an electrochemical
sensor is provided having co-functionalized working electrodes.
Co-functionalization can permit the use of hydrocarbon molecules as
space fillers to cover sites on a silicon hydride (--Si--H) surface
that have not been occupied by redox active moieties, such as
anthracene and/or ferrocene. The hydrocarbon molecules can block
(or passivate) such sites, thereby preventing such sites from
interacting with anions and cations during operation of the
electrochemical sensor, which can provide for improved device
performance.
[0373] In an embodiment, an electrochemical sensor comprises one or
more working electrodes, a counter electrode and a reference
electrode, wherein at least one of the one or more working
electrodes is co-functionalized with redox-active moieties and
hydrocarbon molecules. In an embodiment, an electrochemical sensor
includes a working electrode, a counter electrode and a reference
electrode. The working electrode can include a mixed monolayer of
redox active species, such as anthracene and ferrocene, and
hydrocarbon molecules, such as alkanes, alkenes, or alkynes. In an
embodiment, the hydrocarbon is a long-chain hydrocarbon. In another
embodiment, the hydrocarbon comprises a chain of 2 or more, 3 or
more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or
more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more,
15 or more carbon atoms. In an embodiment, the hydrocarbon
molecules can include a decyne having the following structural
formula:
##STR00002##
[0374] In an embodiment, a working electrode comprises a
submonolayer coverage redox-active species, with the remaining
sites (i.e., S--H sites) covered by a hydrocarbon, such as decyne.
In another embodiment, a working electrode can include a mixed
layer of ferrocene and a hydrocarbon, such as decyne. In another
embodiment, a working electrode can include a mixed layer of
anthracene and a hydrocarbon, such as decyne. In another
embodiment, a working electrode can include a mixed layer of
ferrocene, anthracene, and a hydrocarbon, such as decyne.
Single-Use or Disposable Electrochemical Sensors
[0375] In another aspect of the invention, single-use and/or
disposable electrochemical sensors are provided. In an embodiment,
electrochemical sensors are provided that are suitable for single
use applications, disposable applications, or both single-use and
disposable applications.
[0376] In an embodiment, a disposable electrochemical sensor is
provided. In another embodiment, a single-use electrochemical
sensor is provided. In another embodiment, a disposable and
single-use electrochemical sensor is provided.
[0377] In an embodiment, a single-use electrochemical sensor
comprises an onboard energy storage device. Such an energy storage
device can be configured to store electrical potential energy
(e.g., in the form of electrons in excited states) and discharge
upon use of the electrochemical sensor. Energy storage devices can
be selected from batteries, capacitors, or photovoltaic modules. In
an embodiment, an electrochemical sensor comprises a battery, such
as, e.g., a lithium ion battery or nickel metal cadmium battery. In
another embodiment, an electrochemical sensor comprises a
capacitor. In another embodiment, an electrochemical sensor
comprises a battery electrically coupled to a capacitor.
[0378] In another embodiment, an electrochemical sensor is provided
having a replaceable energy storage device, such as a replaceable
battery. In such a case, the electrochemical device can be used for
a duration determined by the lifetime of the energy storage device.
The energy storage device can be subsequently changed, thereby
permitting further use of the electrochemical sensor. In another
embodiment, an electrochemical sensor having a rechargeable energy
storage device is provided. The rechargeable energy storage device
can include a rechargeable battery.
[0379] In an embodiment, an electrochemical sensor, such as any
electrochemical sensor provided herein (e.g., a pH sensor), can
include an energy storage device configured to provide charge to
the electrochemical sensor. In another embodiment, an
electrochemical sensor includes an energy storage device configured
to provide charge for up to about 1 day, or 2 days, or 3 days, or 4
days, or 5 days, or 6 days, or 7 days, or 8 days, or 9 days, or 10
days, or 11 days, or 12 days, or 13 days, or 14 days, or 15 days,
or 16 days, or 17 days, or 18 days, or 19 days, or 20 days, or 21
days, or 22 days, or 23 days, or 24 days, or 25 days, or 26 days,
or 27 days, or 28 days, or 29 days, or 30 days, or 35 days, or 40
days, or 45 days, or 50 days, or 100 days, or 150 days, or 200
days, or 300 days, or 400 days, or 500 days, or 1000 days. In
another embodiment, an electrochemical sensor includes an energy
storage device configured to provide charge for at least about 1
day, or 2 days, or 3 days, or 4 days, or 5 days, or 6 days, or 7
days, or 8 days, or 9 days, or 10 days, or 11 days, or 12 days, or
13 days, or 14 days, or 15 days, or 16 days, or 17 days, or 18
days, or 19 days, or 20 days, or 21 days, or 22 days, or 23 days,
or 24 days, or 25 days, or 26 days, or 27 days, or 28 days, or 29
days, or 30 days, or 35 days, or 40 days, or 45 days, or 50 days,
or 100 days, or 150 days, or 200 days, or 300 days, or 400 days, or
500 days, or 1000 days.
[0380] In an embodiment, a semiconductor-based electrochemical
sensor is provided having redox-active moieties. The sensor can
include one redox-active moiety that is sensitive to an analyte
(e.g., hydrogen ions) and another redox-active moiety that is
insensitive to the analyte. The semiconductor-based electrochemical
sensor can further include an on-board energy storage device
configured to provide power to the semiconductor-based
electrochemical sensor. In an embodiment, the energy storage device
can provide power to the semiconductor-based electrochemical sensor
for a time period of at least about 1 second, or 10 seconds, or 30
seconds, or 1 minute, or 2 minutes, or 3 minutes, or 4 minutes, or
5 minutes, or 6 minutes, or 7 minutes, or 8 minutes, or 9 minutes,
or 10 minutes, or 30 minutes, or 1 hour, or 2 hours, or 3 ours, or
4 hours, or 5 hours, or 6 hours, or 7 hours, or 8 hours, or 9
hours, or 10 hours, or 11 hours, or 12 hours, or 1 day, or 2 days,
or 3 days, or 4 days, or 5 days, or 6 days, or 7 days, or 8 days,
or 9 days, or days, or 11 days, or 12 days, or 13 days, or 14 days,
or 15 days, or 16 days, or 17 days, or 18 days, or 19 days, or 20
days, or 21 days, or 22 days, or 23 days, or 24 days, or 25 days,
or 26 days, or 27 days, or 28 days, or 29 days, or 30 days, or 35
days, or 40 days, or 45 days, or 50 days, or 100 days, or 150 days,
or 200 days, or 300 days, or 1 year, or 2 years, or 3 years, or 4
years, or 5 years, or 6 years, or 7 years, or 8 years, or 9 years,
or 10 years, or 15 years, or 20 years.
[0381] In an embodiment, a semiconductor-based electrochemical
sensor is provided that is configured for single-use operation. In
another embodiment, an electrochemical sensor is provided that is
configured to operate, upon first use, for a time period of at
least about 1 second, or 10 seconds, or 30 seconds, or 1 minute, or
2 minutes, or 3 minutes, or 4 minutes, or 5 minutes, or 6 minutes,
or 7 minutes, or 8 minutes, or 9 minutes, or 10 minutes, or 30
minutes, or 1 hour, or 2 hours, or 3 ours, or 4 hours, or 5 hours,
or 6 hours, or 7 hours, or 8 hours, or 9 hours, or 10 hours, or 11
hours, or 12 hours, or 1 day, or 2 days, or 3 days, or 4 days, or 5
days, or 6 days, or 7 days, or 8 days, or 9 days, or 10 days, or 11
days, or 12 days, or 13 days, or 14 days, or 15 days, or 16 days,
or 17 days, or 18 days, or 19 days, or 20 days, or 21 days, or 22
days, or 23 days, or 24 days, or 25 days, or 26 days, or 27 days,
or 28 days, or 29 days, or days, or 35 days, or 40 days, or 45
days, or 50 days, or 100 days, or 150 days, or 200 days, or 300
days, or 1 year, or 2 years, or 3 years, or 4 years, or 5 years, or
6 years, or 7 years, or 8 years, or 9 years, or 10 years, or 15
years, or 20 years. In another embodiment, after first use, the
electrochemical sensor will not function (i.e., the electrochemical
sensor will not function beyond the first use).
[0382] In another embodiment, an electrochemical sensor comprising
one or more working electrodes, a counter electrode and a reference
electrode is disposed in a high density polyethylene support that
can be directly welded to a surface a bag, such as a polyethylene
bag, for single use bag application. Such electrochemical sensors
can be configured for use with cell fermentation, media storage,
buffer preparation and cell storage.
[0383] In an embodiment, a redox-active moiety-containing analyte
sensor is provided that is configured for use in a time period of
about 1 day, or 5 days, or 10 days, or 20 days, or 25 days, or 30
days, or 1 month, or 2 months, or 3 months, or 4 months, or 5
months, or 6 months, or 7 months, or 8 months, or 9 months, or 10
months, or 11 months, or 1 year, or 2 years, or 3 years, or 4
years, or 5 years, or 6 years, or 7 years, or 8 years, or 9 years,
or 10 years. In another embodiment, a redox-active
moiety-containing analyte sensor is provided that is configured for
use in a time period of at least about 1 day, or 5 days, or 10
days, or 20 days, or 25 days, or 30 days, or 1 month, or 2 months,
or 3 months, or 4 months, or 5 months, or 6 months, or 7 months, or
8 months, or 9 months, or 10 months, or 11 months, or 1 year, or 2
years, or 3 years, or 4 years, or 5 years, or 6 years, or 7 years,
or 8 years, or 9 years, or 10 years. Such analyte sensors can be
configured for use within a particular time period without
re-calibration.
[0384] Analyte sensors provided herein can have various
sensitivities. In an embodiment, a redox-active moiety-containing
analyte sensor is provided having a sensitivity between about 5 mV
per pH unit and 200 mV per pH unit, or between about 10 mV per pH
unit and 100 mV per pH unit, or between about 20 mV per pH unit and
60 mV per pH unit.
[0385] Analyte sensors provided herein can have various shelf
lives. In an embodiment, a redox-active moiety-containing analyte
sensor is provided having a shelf life between about 1 month and 20
years, or between about 2 months and 10 years, or between about 3
months and 3 years. Such analyte sensors can function without
re-calibration. In another embodiment, a redox-active
moiety-containing analyte sensor is provided having a shelf life of
at least about 1 month, or 2 months, or 3 months, or 4 months, or 5
months, or 6 months, or 7 months, or 8 months, or 9 months, or 10
months, or 11 months, or 1 year, or 2 years, or 3 years, or 4
years, or 5 years, or 6 years, or 7 years, or 8 years, or 9 years,
or 10 years.
[0386] Analyte sensors provided herein can have various accuracies
following removal from storage. In an embodiment, a sensor for
detecting an analyte is provided having an accuracy to within 0.1%
(pH units) while in use or storage for at least 2 years. In another
embodiment, a sensor for detecting an analyte is provided having an
accuracy to within 0.1% (pH units) while in use or storage for at
least 4 years. In another embodiment, a sensor for detecting an
analyte is provided having an accuracy to within 0.1% (pH units)
while in use or storage for at least 8 years. In another
embodiment, a sensor for detecting an analyte is provided having an
accuracy to within 0.1% (pH units) while in use or storage for at
least 10 years. In some embodiments, sensors can retain their
accuracy while in storage without the need for calibration of
re-calibration. Such sensors can include redox-active moieties, as
described herein.
[0387] In an embodiment, a sensor for detecting an analyte is
provided having an accuracy of plus or minus 0.0001 pH units, or
0.0002 pH units, or 0.0003 pH units, or 0.0004 pH units, or 0.0005
pH units, or 0.0006 pH units, or 0.0007 pH units, or 0.0008 pH
units, or 0.0009 pH units, or 0.001 pH units, or 0.002 pH units, or
0.003 pH units, or 0.004 pH units, or 0.005 pH units, or 0.006 pH
units, or 0.007 pH units, or 0.008 pH units, or 0.009 pH units, or
0.01 pH units, or 0.02 pH units, or 0.03 pH units, or 0.04 pH
units, or 0.05 pH units, or 0.06 pH units, or 0.07 pH units, or
0.08 pH units, or 0.09 pH units, or 0.1 pH units. Such a sensor can
include redox-active moieties, as described herein.
[0388] In an embodiment, a sensor for detecting an analyte is
provided having an accuracy to within 0.001 pH units while in use
or storage for at least 2 years. In another embodiment, a sensor
for detecting an analyte is provided having an accuracy to within
0.001 pH units while in use or storage for at least 4 years. In
another embodiment, a sensor for detecting an analyte is provided
having an accuracy to within 0.001 pH units while in use or storage
for at least 8 years. In another embodiment, a sensor for detecting
an analyte is provided having an accuracy to within 0.001 pH units
while in use or storage for at least 10 years. In some embodiments,
sensors can retain their accuracy while in storage without the need
for calibration of re-calibration. Such sensors can include
redox-active moieties, as described herein.
[0389] In an embodiment, a sensor for detecting an analyte is
provided having an accuracy to within 0.0001, or 0.001, or 0.01, or
0.1 pH units while in use or storage for at least 1 year. In
another embodiment, a sensor for detecting an analyte is provided
having an accuracy to within 0.0001, or 0.001, or 0.01, or 0.1 pH
units while in use or storage for at least 2 years. In another
embodiment, a sensor for detecting an analyte is provided having an
accuracy to within 0.0001, or 0.001, or 0.01, or 0.1 pH units while
in use or storage for at least 4 years. In another embodiment, a
sensor for detecting an analyte is provided having an accuracy to
within 0.0001, or 0.001, or 0.01, or 0.1 pH units while in use or
storage for at least 8 years. In another embodiment, a sensor for
detecting an analyte is provided having an accuracy to within
0.0001, or 0.001, or 0.01, or 0.1 pH units while in use or storage
for at least 10 years. Such sensors can include redox-active
moieties, as described herein.
[0390] In an embodiment, a redox-active moiety-containing analyte
sensor is provided having a shelf life of at least about 1 month,
or 2 months, or 3 months, or 4 months, or 5 months, or 6 months, or
7 months, or 8 months, or 9 months, or 10 months, or 11 months, or
1 year, or 2 years, or 3 years, or 4 years, or 5 years, or 6 years,
or 7 years, or 8 years, or 9 years, or 10 years, and an accuracy to
within about 0.01%, or 0.02%, or 0.03%, or 0.04%, or 0.05%, or
0.1%, or 0.15%, or 0.2%. In another embodiment, a redox-active
moiety-containing analyte sensor is provided having a shelf life of
at least about 1 month, or 2 months, or 3 months, or 4 months, or 5
months, or 6 months, or 7 months, or 8 months, or 9 months, or
months, or 11 months, or 1 year, or 2 years, or 3 years, or 4
years, or 5 years, or 6 years, or 7 years, or 8 years, or 9 years,
or 10 years, and an accuracy to within about 0.0001 pH units, or
0.0002 pH units, or 0.0003 pH units, or 0.0004 pH units, or 0.0005
pH units, or 0.0006 pH units, or 0.0007 pH units, or 0.0008 pH
units, or 0.0009 pH units, or 0.001 pH units, or 0.002 pH units, or
0.003 pH units, or 0.004 pH units, or 0.005 pH units, or 0.006 pH
units, or 0.007 pH units, or 0.008 pH units, or 0.009 pH units, or
0.01 pH units, or 0.02 pH units, or 0.03 pH units, or 0.04 pH
units, or 0.05 pH units, or 0.06 pH units, or 0.07 pH units, or
0.08 pH units, or 0.09 pH units, or 0.1 pH units. Such sensors can
include redox-active moieties, as described herein.
[0391] In an embodiment, a redox-active moiety-containing analyte
sensor is provided that can be used to measure the presence or
absence (or concentration) or an analyte, without re-calibration,
for at least about 1 day, or 1 week, or 2 weeks, or 1 month, or 2
months, or 3 months, or 4 months, or 5 months, or 6 months, or 7
months, or 8 months, or 9 months, or 10 months, or 11 months, or 1
year, or 2 years, or 3 years, or 4 years, or 5 years, or 6 years,
or 7 years, or 8 years, or 9 years, or 10 years, at an accuracy to
within about 0.01%, or 0.02%, or 0.03%, or 0.04%, or 0.05%, or
0.1%, or 0.15%, or 0.2%. In some cases, the accuracy is to within
about 10%, or 5%, or 1%, or 0.1%, or 0.01%, or 0.01%, or 0.001%. In
another embodiment, a redox-active moiety-containing analyte sensor
is provided that can be used to measure the presence or absence (or
concentration) of an analyte, without re-calibration, for at least
about 1 day, or 1 week, or 2 weeks, or 1 month, or 2 months, or 3
months, or 4 months, or 5 months, or 6 months, or 7 months, or 8
months, or 9 months, or 10 months, or 11 months, or 1 year, or 2
years, or 3 years, or 4 years, or 5 years, or 6 years, or 7 years,
or 8 years, or 9 years, or 10 years, and an accuracy to within
about 0.0001 pH units, or 0.0002 pH units, or 0.0003 pH units, or
0.0004 pH units, or 0.0005 pH units, or 0.0006 pH units, or 0.0007
pH units, or 0.0008 pH units, or 0.0009 pH units, or 0.001 pH
units, or 0.002 pH units, or 0.003 pH units, or 0.004 pH units, or
0.005 pH units, or 0.006 pH units, or 0.007 pH units, or 0.008 pH
units, or 0.009 pH units, or 0.01 pH units, or 0.02 pH units, or
0.03 pH units, or 0.04 pH units, or 0.05 pH units, or 0.06 pH
units, or 0.07 pH units, or 0.08 pH units, or 0.09 pH units, or 0.1
pH units. Such sensors can include redox-active moieties, as
described herein.
[0392] In some embodiments, a method for detecting the presence or
absence of an analyte, comprises bringing an analyte sensor in
contact with a sample, said analyte sensor having an electrode
having immobilized thereon a redox-active moiety, wherein the
redox-active moiety exhibits an oxidation potential and/or a
reduction potential that is sensitive to the presence of said
analyte. Next, with the aid of the analyte sensor, the analyte is
detected to an accuracy of within about 20% without re-calibration
for a period of at least about 1 day. In some cases the accuracy
can be to within about 10%, or 5%, or 1%, or 0.1%, or 0.01%, or
0.01%, or 0.001%. In some embodiments, the analyte sensor can
detect the presence or absence of the analyte without
re-calibration for a time period of at least about 1 day, or 1
week, or 2 weeks, or 1 month, or 2 months, or 3 months, or 4
months, or 5 months, or 6 months, or 7 months, or 8 months, or 9
months, or 10 months, or 11 months, or 1 year, or 2 years, or 3
years, or 4 years, or 5 years, or 6 years, or 7 years, or 8 years,
or 9 years, or 10 years.
Electrochemical Sensors with Multiple Sensor Modules
[0393] In another aspect of the invention, electrochemical sensors
are provided having a plurality of sensor modules, each sensor
module configured to measure an analyte of interest. In an
embodiment, an electrochemical sensor comprises at least 2, or 3,
or 4, or 5, or 6, or 7, or 8, or 9, or 10, or 15, or 20, or 25, or
30, or 35, or 40, or 45, or 50, or 100 modules. In another
embodiment, an electrochemical sensor comprises a plurality of
modules. The modules can be configured to detect (or measure) the
presence of the same analyte (e.g., H+) or different analytes
(e.g., H+ and O.sub.2).
[0394] With reference to FIG. 22A, an electrochemical sensor having
three modules, wherein each module is configured to sense an
analyte of interest, is illustrated, in accordance with an
embodiment of the invention. The electrochemical sensor comprises a
probe body that can be cylindrical in shape. FIG. 22B is an
enlarged view of a portion of the electrochemical sensor of FIG.
22A, illustrating three modules configured to sense (or measure the
presence of) one or more analytes of interest. Each of the three
modules can be configured to sense the same analyte (e.g., H+) or
different analytes. For example, a first module can be configured
to sense (and measure the concentration of) H+, a second module can
be configured to sense O.sub.2, and a third module can be
configured to sense NH.sub.3. Such "multi-modal" sensor can be
advantageous for use in chemical reactors and bioreactors, where
the measurement of multiple analytes is desirable.
Electrochemical Sensors Formed on Printed Circuit Boards
[0395] In another aspect of the invention, electrochemical sensors
formed on circuit boards are provided. With reference to FIG. 23,
In an embodiment, a printed circuit board is provided. A dam can be
placed on a field that fits around the printed circuit board. The
field provides a support surface that can be free of contaminants.
The dam can constrain the area between the field and the printed
circuit board. A filler material ("filler") can be injected through
holes in the printed circuit board. The filler can hold the
electrodes in place and exclude fluids from touching the back side
of the electrodes and the traces of the printed circuit board. The
filler can then be cured. Following curing, the sensor can be
removed from the field and mounted into an appropriate holder, such
as a probe body (see FIG. 22A).
[0396] Sensors formed on printed circuit boards can be mounted in a
wide array of form factors. For example, they can be placed in high
density polyethylene hubs and welded into disposable polyethylene
containers. As another example, they can be mounted within tube
walls or at the entrance and egress of columns Such tubes walls or
columns can be configured for use with containers having dimensions
(e.g., diameters, lengths) configured for use with glass probe
electrochemical (e.g., pH measurement) systems.
[0397] With reference to FIG. 24, In an embodiment, a sensor formed
on a printed circuit board can be mounted on a head assembly. Such
head assembly can be configured to be mounted on a tube assembly or
column, such as the tip of a tube assembly or column. In another
embodiment, a head assembly can be removably mounted to a tube
assembly or column. In another embodiment, a head assembly can be
irremovably mounted to a tube assembly or column. In another
embodiment, a head assembly can be mounted to a top portion of a
tube or column for insertion into a solution having an analyte of
interest. A protective material is applied around the head assembly
to prevent the solution from entering a back portion of the head
assembly. In an embodiment, the protective material includes one or
more of polyurethane and an epoxy (e.g., polyexpoxide).
[0398] With continued reference to FIG. 24, the sensor includes a
surface configured to come in contact with a solution having one or
more analytes. The sensor includes a counter electrode (top) and a
reference electrode (center). The sensor further includes working
electrodes (bottom left and bottom right). One working electrode
comprises a semiconductor surface that has immobilized thereon a
redox active moiety whose oxidation potential and/or reduction
potential is sensitive to the presence of the analyte, and a second
working electrode comprises redox active moiety whose oxidation
potential and/or reduction potential is insensitive to the presence
of the analyte (see above).
Electrochemical Sensors for Use with Containers and Flow-Through
Containers
[0399] In another aspect of the invention, an electrochemical
sensor, such as any sensor provided herein can be used with
containers and reactors such as disposable containers and reactors.
In an embodiment, an electrochemical sensor is provided for use
with a disposable flow-through container. In another embodiment, an
electrochemical sensor is provided for use with a disposable
reactor. In another embodiment, an electrochemical sensor is
provided for use with a disposable plug flow reactor. In another
embodiment, an electrochemical sensor is provided for use with a
disposable continuous stirred tank reactor (CSTR).
[0400] In some embodiments, any electrochemical sensor ("sensor")
provided herein can be configured for use with a container,
including a flexible container. In an embodiment, a sensor is
provided configured for use with a disposable flexible container.
Such flexible container can have walls formed of a polymeric
material.
[0401] With reference to FIG. 25A, an electrochemical sensor is
shown mounted on a wall of a container, in accordance with an
embodiment of the invention. The container can be a disposable
container. The electrochemical sensor comprises a printed circuit
board having thereon a counter electrode, a reference electrode and
one or more working electrode (one working electrode illustrated in
the cross-sectional cut-away). The printed circuit board brings the
electrodes in electrical communication with connectors, which can
be used to interface with a computer system or device for use with
the electrochemical sensor. The electrochemical sensor is mounted
on a holder flange, which is separated from the printed circuit
board through a separation member, such as a damn-to-holder ring
("O-Ring", as illustrated). The probe holder can seal to the dam.
The flange of the holder can be substantially thin so that it can
be welded to a container, such as a bag. In an embodiment, a
portion of the container is mounted to the holder flange with the
aid of an adhesive, such as, for example, one or more of
polyurethane and an epoxy (e.g., polyepoxide). In another
embodiment, a portion of the container is welded to the holder
flange. In another embodiment, the electrochemical sensor can be
sterilized, such as with the aid of sterilizing chemicals,
ultraviolet light irradiation, or plasma treatment.
[0402] With reference to FIG. 25B, an electrochemical sensor is
shown formed on a printer circuit board (PCB). The electrochemical
sensor further includes a connector for providing a connection
between the electrochemical sensor and a reader or other
electronics unit, such as a voltage sync (see FIGS. 27A and 27B).
The electrodes of the electrochemical sensor are disposed at a
bottom portion of the PCB. In an embodiment, the electrodes of the
electrochemical sensor, including working electrodes, a counter
electrode and reference electrode, are formed in the PCB.
[0403] In an embodiment, the PCB can be formed of a military
standard PCB board (e.g., FR4 substrate) or any PC board
constructed from a non-conductive material, with conductive traces.
The boards can be multilayered.
[0404] In various embodiments, solutions to seal or hermetically
seal (also "sealing solutions" herein) sensor components are
provided. In an embodiment, sealing solutions can comply with
biological manufacturing standards, such as reducing, if not
eliminating, outgassing, and avoiding the use of environmentally
hazardous or toxic materials. In an embodiment, a sealing solution
is provided to a sensor through the potting technique, or the
application of a liquid adhesive to prevent water and other liquids
from entering the sensor and causing electrical short circuits. In
an embodiment, a PCB having sensor components is mounted to a tube.
An interior portion of the tube is sealed from the external
environment with the aid of a sealing solution around the PCB.
[0405] In another aspect of the invention, electrochemical sensors
are provided for use with fluid flow channels, such as flow-through
tubes or pipes. With reference to FIG. 26, an electrochemical
sensor, such as the sensor discussed in the context of FIG. 25A, is
mounted in a chamber of a flow-through tube. The chamber can be
configured for mating with a flange portion of the electrochemical
sensor. In an embodiment, the electrochemical sensor can be welded
to the chamber. In another embodiment, the electrochemical sensor
can be attached to the chamber with the aid of an epoxy, for
example. In another embodiment, the electrochemical sensor can be
removably attached to the chamber. In another embodiment, the
electrochemical sensor can be irremovably attached to the
chamber.
Electrochemical Sensor Electrical Components
[0406] In some embodiments, the sensor or holder can have on-board
electronic memory, such as Electrically Erasable Programmable
Read-Only Memory ("EEPROM"). The EEPROM can hold or store various
items (or information) that can pair the electrochemical sensor
with drive electronics. In an embodiment, the EEPROM can hold a
serial number of the electrochemical sensor. In another embodiment,
probe calibration parameters (e.g., probe calibration constants)
can be held in the memory of the EEPROM. In another embodiment, the
algorithm details of the electrochemical sensor can be held in the
EEPROM.
[0407] In an embodiment, the EEPROM is provided on a printed
circuit board having the various electrodes of the electrochemical
sensor (e.g., pH sensor). In another embodiment, the EEPROM is
provided adjacent a printed circuit board or other device having
the various electrodes of the electrochemical sensor.
[0408] An electrochemical sensor comprising an on-board EEPROM can
have various advantages and benefits over other electrochemical
sensors. For example, an electrochemical sensor having an on-board
EEPROM can be electrically optimized for different uses or markets.
As an example a probe can be optimized for pH near pH 7 and only
scan for a small region around pH 7. An identical probe could be
optimized for pH 4 and only scan an area around pH 4. A sensor
optimized for pH 4 measurement and a sensor optimized for pH 7
measurements can be similar, if not identical in form and function,
different only the algorithm provided in the EEPROM of each
sensor.
[0409] An EEPROM on-board an electrochemical sensor can save or
hold one or more types of information. In an embodiment, the EEPROM
can hold a counter for measuring (and storing) the length time an
electrochemical device has been used. In another embodiment, if an
electrochemical sensor is configured for use over a certain time
period, the counter can enable the electrochemical sensor to stop
functioning after the time period has been reached. The EEPROM can
be used in conjunction with (or to facilitate) single-use
functionality, as described in certain embodiments (see above). For
example, if the electrochemical sensor is configured for use over
the period of 28 days, the electrochemical sensor can stop
functioning once the 28-day time period has elapsed. In another
embodiment, the counter can enable a user to know how much time the
user has remaining to use the sensor. In an embodiment, as the
sensor is used, a write once area in the EEPROM can be incremented,
and when this area of the EEPROM is filled, it will signal that the
useful life of the sensor is finished, and the sensor will
terminate use. In another embodiment, once the useful life of the
sensor has finished, the user may be prevented from using the
sensor.
[0410] In another embodiment, the EEPROM can transmit its serial
number or other identifying information, in addition to the time
remaining for the electrochemical sensor, to a data recorder that
can monitor one or more sensors in a user's operation.
[0411] Alternatively, sensor calibration parameters can be stored
on a remote computer system (e.g., the "cloud"), such as a server,
and transmitted to a system in electrical communication with the
sensor.
[0412] In some situations, a sensor can include an identification
member, such as radio-frequency identification (RFID) tag, to
enable a system communicating with the sensor to identify the
sensor and determine, for example, any calibration parameters that
may be required by the sensor. The identification member can be on
a body or housing of the sensor. Calibration parameters may be
specific to a particular application (e.g., pH measurements) of the
sensor. The identification member can enable the system to provide
proper calibration parameters, as may be required for a particular
sensor application.
Electronics Components for Integration into Third-Party Systems
[0413] In another aspect of the invention, an electronics component
is provided for emulating various features of current third-party
or traditional electrochemical sensors. Such electronics components
can advantageously permit the integration of electrochemical
sensors provided herein into various current or traditional
systems, such as glass probe systems.
[0414] Many laboratories and manufacturing facilities have existing
third party pH meters, either stand alone or within digital control
units. In some embodiments, an electronics unit is provided for
integration into existing third party pH meters and systems, such
as analog pH meters and systems. In an embodiment, the electronics
unit can do one or more of emulating the functionality of a
potentiostat, providing power the electrochemical sensor, and
communicating with a third party pH meter to provide the user with
a direct pH readout.
[0415] In an embodiment, the electronics unit enables communication
between a third-party reader (or meter) and any probe provided
herein. The electronics unit can further provide power to the
probe. In an embodiment, the electronics unit includes a power
input for providing power to one or more of the electronics unit
and a probe in electrical communication with the electronics unit,
an input from the probe, and an output to a reader.
[0416] With reference to FIG. 27A, an electronic unit is shown, in
accordance with an embodiment of the invention. The electronic
unit, which is illustrated as an electrical box, can permit
integration of electrochemical sensors provided herein into
systems, including electronics components, configured for use with
types of electrochemical systems. With continued reference to FIG.
27A, the electrical box can be connected to a sensor on one side
and a traditional readout unit or device on the other side. In at
least some embodiments, electrochemical sensors are provided that
use microprocessors to control transmitter electronics to scan the
sensor appropriately. The electronics component evaluates the
sensor signals and determines the appropriate sensor value (pH
value in this case). The electronics component then takes the
sensor value and computes the appropriate output voltage or current
to emulate the traditional sensor. The electronics component will
then transmit the appropriate output voltage to a system or device
configured for use with a traditional sensor, such as a glass probe
sensor.
[0417] In some embodiments, the electronic unit of FIG. 27A can
transform a digital signal into a signal that is readable by a
third-party system, such as a system configured for use with a
glass electrode. In an embodiment, the electronic unit can emulate
the glass electrode output. The electronic unit can advantageously
make sensors provided herein compatible with all or substantially
all third party electrochemical sensors, including pH sensors.
[0418] In an embodiment, an electronics unit is provided for
integrating semiconductor-based electrochemical sensors, such as
those provided herein, in devices configured for use with
traditional probes, such as glass probes (e.g., glass pH probes).
In another embodiment, an electronics unit is provided that is a
digital to analog converter. Such an electronics unit can be
configured to enable electrochemical sensors of embodiments to be
used with traditional or current analog probe systems.
[0419] Traditional pH meters can have complex calibration systems
for setting the voltage slope and offset of a glass pH probe. In an
embodiment, a voltage sync is provided that resets a third-party pH
meter. With reference to FIG. 27A, the electronics unit includes
three buttons to permit the electronics unit to emulate the three
point calibration for certain probe systems, which is standard
prior to data collection with an analog pH meter. The electronics
unit of FIG. 27A includes three buttons, each button configured to
enable the electronic unit to output to an electronic system (for
use with a traditional prove) a voltage value that is recognizable
by the electronic system. For example, the button marked pH 7
(middle button) can sets the voltage output of the transmitter to
about zero volts. The button marked pH 4 (left button) can set the
voltage to about +0.177 V (or any other voltage corresponding to pH
4 for the traditional electronics system). The button marked pH 10
(right button) can set the voltage to about -0.177 V (or any other
voltage corresponding to pH 10 for the traditional electronics
system). This way a pH meter that was calibrated to a particular
glass pH probe can be returned to the ideal slope and offset.
[0420] With continued reference to FIG. 27A, the electronic unit
includes one cable or corrector for interfacing with
electrochemical sensors provided herein (left cable), another
connector for interfacing with a traditional electronics unit
(right-top cable), such as an electronics unit configured for use
with glass probe electrochemical sensors (e.g., pH sensors), and
another connector (right-bottom cable) for providing power to the
electrochemical sensor.
[0421] While in certain embodiments a stand-alone electronics units
is provided, in other embodiments, the electronics units can be
included on-board the printed circuit board having the
electrochemical sensor (see FIGS. 23 and 24).
[0422] FIG. 27B illustrates the electronic unit of FIG. 27A
attached to (or electrical communication with) a probe, in
accordance with an embodiment of the invention. The probe can be
any probe described herein, such as, for example, the probe of FIG.
1B. The probe comprises a cylindrical body and a head portion. The
head portion can include one or more working electrodes, a
reference electrode and a counter electrode, as described herein.
The head portion is configured to come in contact with a solution
having one or more analytes of interest, such as, e.g., hydrogen
ions (H+).
[0423] FIG. 27C shows a system having a probe attached to an
electronics unit, such as the electronics unit of FIG. 27A, in
accordance with an embodiment of the invention. The system further
includes a reader (or meter) for providing sensor measurements,
such as pH measurements. In an embodiment, the reader is a digital
reader. In another embodiment, the electronic unit can be
configured for enabling communication between a reader, such as a
third-party reader, and the probe. The third-party reader can be
configured for use with glass probe electrochemical sensors, such
as glass probe pH sensors.
[0424] In an embodiment, the electronics unit can be configured for
wireless communication with a probe. In another embodiment, the
electronics unit can include one or more of a WiFi transmitter, a
Bluetooth transmitter, a radiofrequency transmitter and an infrared
(IR) transmitter for communicating with the probe.
Electronics Components for Communicating with External Devices
[0425] Another aspect of the invention provides an electrochemical
sensor, such as an electrochemical sensor on a printed circuit
board, which includes a transmitter for wirelessly transmitting
information to a system configured to collect information from the
electrochemical sensor. The transmitter can be reduced in size so
that it can be included in the sensor (or probe). The transmitter
can be configured to interface with a receiver on a system
configured to communicate with the electrochemical sensor. In an
embodiment, the electrochemical sensor can be configured for WiFi
transmission. In another embodiment, the electrochemical sensor can
be configured for Bluetooth transmission. In another embodiment,
the electrochemical sensor can be configured for radiofrequency
(RF) transmission. In another embodiment, the electrochemical
sensor can be configured for infrared transmission. In another
embodiment, the electrochemical sensor can be configured for
inductive transmission (i.e., inductively coupling). In another
embodiment, the electrochemical sensor can be configured for
optical (e.g., fiber optic) transmission.
[0426] In some embodiments, a transmitter is provided that is
powered by an energy storage device, such as a battery (e.g.,
lithium ion battery) or a photovoltaic (solar) cell. The energy
storage device can be configured to provide power to the
transmitter over the life of the electrochemical sensor. In an
embodiment, the energy storage device is configured to provide
power to both the transmitter and the electrochemical sensor, such
as any electrochemical sensor provided herein. In another
embodiment, the transmitter can be an on-board transmitter. For
example, the electrochemical sensor of FIG. 24, configured to be
mounted at a tip of a probe assembly or shaft, can include a
transmitter. The transmitter can include electronics that can
provide a wireless link, such as a digital link (e.g., WiFi,
Bluetooth) to a computer system. This link can be done with WiFi,
Bluetooth or other radio communications protocols, such as
radiofrequency (RF) protocols.
[0427] An electrochemical sensor configured for wireless
transmission can be used in numerous contexts. In an embodiment, an
electrochemical sensor configured for wireless transmission can be
configure to transmit sensor data from remote and hard-to-reach
locations, such as providing data from inside oil wells, the human
body (e.g., providing the concentrations of various analytes, such
as pH) and high pressure settings, such as chemical reactors.
[0428] In another embodiment, an electronics component is provided
that can be configured to provide a digital signal that can be read
using a computing device, such as a personal computer (PC) or a
mobile electronics device, such as an Apple.RTM. iPad.RTM. or
iPod.RTM., an Android.RTM.-enabled device, a Smart Phone, a
netbook, a laptop, a tablet PC, or a slate PC.
[0429] In an embodiment, a graphical user interface ("GUI") can
provide a user a sensor reading, such as at a fixed point in time
or as a function of time. In an embodiment, the GUI can provide a
user a sensor reading (e.g., pH reading) at least every 0.1
seconds, or 0.2 seconds, or 0.3 seconds, or 0.4 seconds, or 0.5
seconds, or 0.6 seconds, or 0.7 seconds, or 0.8 seconds, or 0.9
seconds, or 1 second, or 1.1 seconds, or 1.2 seconds, or 1.3
seconds, or 1.4 seconds, or 1.5 seconds, or 1.6 seconds, or 1.7
seconds, or 1.8 seconds, or 1.9 seconds, or 2 seconds, or 2.1
seconds, or 2.2 seconds, or 2.3 seconds, or 2.4 seconds, or 2.5
seconds, or 2.6 seconds, or 2.7 seconds, or 2.8 seconds, or 2.9
seconds, or 3 seconds, or 3.1 seconds, or 3.2 seconds, or 3.3
seconds, or 3.4 seconds, or 3.5 seconds, or 3.6 seconds, or 3.7
seconds, or 3.8 seconds, or 3.9 seconds, or 4 seconds, or 4.1
seconds, or 4.2 seconds, or 4.3 seconds, or 4.4 seconds, or 4.5
seconds, or 4.6 seconds, or 4.7 seconds, or 4.8 seconds, or 4.9
seconds, or 5 seconds, or 5.1 seconds, or 5.2 seconds, or 5.3
seconds, or 5.4 seconds, or 5.5 seconds, or 5.6 seconds, or 5.7
seconds, or 5.8 seconds, or 5.9 seconds, or 6 seconds, or 6.1
seconds, or 6.2 seconds, or 6.3 seconds, or 6.4 seconds, or 6.5
seconds, or 6.6 seconds, or 6.7 seconds, or 6.8 seconds, or 6.9
seconds, or 7 seconds, or 7.1 seconds, or 7.2 seconds, or 7.3
seconds, or 7.4 seconds, or 7.5 seconds, or 7.6 seconds, or 7.7
seconds, or 7.8 seconds, or 7.9 seconds, or 8 seconds, or 8.1
seconds, or 8.2 seconds, or 8.3 seconds, or 8.4 seconds, or 8.5
seconds, or 8.6 seconds, or 8.7 seconds, or 8.8 seconds, or 8.9
seconds, or 9 seconds, or 9.1 seconds, or 9.2 seconds, or 9.3
seconds, or 9.4 seconds, or 9.5 seconds, or 9.6 seconds, or 9.7
seconds, or 9.8 seconds, or 9.9 seconds, or 10 seconds, or 10.1
seconds, or 10.2 seconds, or 10.3 seconds, or 10.4 seconds, or 10.5
seconds, or 10.6 seconds, or 10.7 seconds, or 10.8 seconds, or 10.9
seconds, or 11 seconds, or 11.1 seconds, or 11.2 seconds, or 11.3
seconds, or 11.4 seconds, or 11.5 seconds, or 11.6 seconds, or 11.7
seconds, or 11.8 seconds, or 11.9 seconds, or 12 seconds, or 12.1
seconds, or 12.2 seconds, or 12.3 seconds, or 12.4 seconds, or 12.5
seconds, or 12.6 seconds, or 12.7 seconds, or 12.8 seconds, or 12.9
seconds, or 13 seconds, or 13.1 seconds, or 13.2 seconds, or 13.3
seconds, or 13.4 seconds, or 13.5 seconds, or 13.6 seconds, or 13.7
seconds, or 13.8 seconds, or 13.9 seconds, or 14 seconds, or 14.1
seconds, or 14.2 seconds, or 14.3 seconds, or 14.4 seconds, or 14.5
seconds, or 14.6 seconds, or 14.7 seconds, or 14.8 seconds, or 14.9
seconds, or 15 seconds, or 15.1 seconds, or 15.2 seconds, or 15.3
seconds, or 15.4 seconds, or 15.5 seconds, or 15.6 seconds, or 15.7
seconds, or 15.8 seconds, or 15.9 seconds, or 16 seconds, or 16.1
seconds, or 16.2 seconds, or 16.3 seconds, or 16.4 seconds, or 16.5
seconds, or 16.6 seconds, or 16.7 seconds, or 16.8 seconds, or 16.9
seconds, or 17 seconds, or 17.1 seconds, or 17.2 seconds, or 17.3
seconds, or 17.4 seconds, or 17.5 seconds, or 17.6 seconds, or 17.7
seconds, or 17.8 seconds, or 17.9 seconds, or 18 seconds, or 18.1
seconds, or 18.2 seconds, or 18.3 seconds, or 18.4 seconds, or 18.5
seconds, or 18.6 seconds, or 18.7 seconds, or 18.8 seconds, or 18.9
seconds, or 19 seconds, or 19.1 seconds, or 19.2 seconds, or 19.3
seconds, or 19.4 seconds, or 19.5 seconds, or 19.6 seconds, or 19.7
seconds, or 19.8 seconds, or 19.9 seconds, or 20 seconds or 20.1
seconds, or 20.2 seconds, or 20.3 seconds, or 20.4 seconds, or 20.5
seconds, or 20.6 seconds, or 20.7 seconds, or 20.8 seconds, or 20.9
seconds, or 21 seconds, or 21.1 seconds, or 21.2 seconds, or 21.3
seconds, or 21.4 seconds, or 21.5 seconds, or 21.6 seconds, or 21.7
seconds, or 21.8 seconds, or 21.9 seconds, or 22 seconds, or 22.1
seconds, or 22.2 seconds, or 22.3 seconds, or 22.4 seconds, or 22.5
seconds, or 22.6 seconds, or 22.7 seconds, or 22.8 seconds, or 22.9
seconds, or 23 seconds, or 23.1 seconds, or 23.2 seconds, or 23.3
seconds, or 23.4 seconds, or 23.5 seconds, or 23.6 seconds, or 23.7
seconds, or 23.8 seconds, or 23.9 seconds, or 24 seconds, or 24.1
seconds, or 24.2 seconds, or 24.3 seconds, or 24.4 seconds, or 24.5
seconds, or 24.6 seconds, or 24.7 seconds, or 24.8 seconds, or 24.9
seconds, or 25 seconds, or 25.1 seconds, or 25.2 seconds, or 25.3
seconds, or 25.4 seconds, or 25.5 seconds, or 25.6 seconds, or 25.7
seconds, or 25.8 seconds, or 25.9 seconds, or 26 seconds, or 26.1
seconds, or 26.2 seconds, or 26.3 seconds, or 26.4 seconds, or 26.5
seconds, or 26.6 seconds, or 26.7 seconds, or 26.8 seconds, or 26.9
seconds, or 27 seconds, or 27.1 seconds, or 27.2 seconds, or 27.3
seconds, or 27.4 seconds, or 27.5 seconds, or 27.6 seconds, or 27.7
seconds, or 27.8 seconds, or 27.9 seconds, or 28 seconds, or 28.1
seconds, or 28.2 seconds, or 28.3 seconds, or 28.4 seconds, or 28.5
seconds, or 28.6 seconds, or 28.7 seconds, or 28.8 seconds, or 28.9
seconds, or 29 seconds, or 29.1 seconds, or 29.2 seconds, or 29.3
seconds, or 29.4 seconds, or 29.5 seconds, or 29.6 seconds, or 29.7
seconds, or 29.8 seconds, or 29.9 seconds, or 30 seconds, or 31
seconds, or 32 seconds, or 33 seconds, or 34 seconds, or 35
seconds, or 36 seconds, or 37 seconds, or 38 seconds, or 38
seconds, or 40 seconds, or 45 seconds, or 50 seconds, or 55
seconds, or 1 minute, or 2 minutes, or 3 minutes, or 4 minutes, or
5 minutes, or 6 minutes, or 7 minutes, or 8 minutes, or 9 minutes,
or 10 minutes, or 20 minutes, or 30 minutes, or 1 hour, or 2 hours,
or 3 hours, or 4 hours, or 5 hours, or 6 hours, or 7 hours, or 8
hours, or 9 hours, or 10 hours, or 11 hours, or 12 hours, or 1 day,
or 2 days, or 3 days, or 4 days, or 5 days.
[0430] In an embodiment, the electrochemical sensor can provide a
reading to a computer system coupled to (or interfaced with) the
electrochemical sensor. Such a reading can be recorded in the
computer's memory with a timestamp that can be assembled into a
date and/or time based graph. The GUI can also record multiple
sensors outputs, such as from a probe having a plurality of
electrochemical sensors (see FIGS. 22A and 22B). The GUI can
compile data from different sensors and provide a user an output
that is calculated from the multiple sensors. In an embodiment,
such output can be an average output, such as an average pH. For
example, if pH measurements are being made, the curve for pH can be
temperature dependent, and the system can correct the pH for
temperature. The system can calculate and provide an input for the
temperature and concentration of hydrogen ions.
[0431] In an embodiment, a sensor is provided having
electrochemical sensors and sensors for measuring the temperature
of a solution or fluid in the solution or fluid. The temperature
can be the temperatures as measured in the vicinity of the sensor.
In an embodiment, the sensor includes one or more thermocouples for
measuring the temperature of a solution adjacent to the sensor.
Such temperature measurement can be used to provide a
temperature-corrected sensor output, such as a pH output corrected
for temperature.
[0432] In some embodiments, a digital meter is provided that can
display the readout from an electrochemical sensor. In an
embodiment, the digital meter can provide real-time data. In
another embodiment, the digital meter can provide date at set
intervals. In such a case, data can be averaged over a certain time
period that can be a function of the intervals in which data is
measured and/or displayed. The digital meter can provide one or
more of current, voltage, impedance, conductivity, temperature,
time, time left in the lifetime of the device (e.g., if the
electrochemical sensor is a time-limited device, see above), area
calculations (e.g., IV area calculations) and maximum peak
positions in an IV curve. A sensor trend can also be displayed. In
an embodiment, the meter can communicate digitally with industrial
digital control units, such as control units in coal fired power
plants, steam boiler water, food and beverage manufacturing
facilities, personal care products, water and wastewater,
refineries, biofuel manufacturing facilities, reactors,
bioreactors, oil well rigs, oil well boars, or nuclear power plants
and the manufacture of radionucleide therapeutics.
Sensors for Insertion into Containers for Use with Glass Probes
[0433] Another aspect of the invention provides a sensor system
comprising a redox-active moiety-containing analyte sensor for
insertion into a container for use with a glass probe analyte
sensor. In an embodiment, the redox-active moiety-containing
analyte sensor comprises one or more redox-active moieties. In
another embodiment, the redox-active moiety-containing analyte
sensor comprises a redox-active moiety that is sensitive to the
presence of an analyte and another redox-active moiety that is
insensitive to the presence of the analyte. In another embodiment,
the redox-active moiety-containing analyte sensor is disposed in a
probe body having a form factor configured for insertion into a
container for use with a glass probe analyte sensor.
[0434] In an embodiment, an electrochemical sensor is provided
having a sensor head, the sensor head having working, counter and
reference electrodes, as described above. The sensor head can be
mounted to a probe body having a shape (size and length) configured
to mate with existing or traditional probe systems, such as glass
probe systems. The electrochemical sensor can be interfaced with a
system used to operate the traditional probe system with the aid of
the electrical box described above in the context of FIG. 27.
[0435] With reference to FIG. 28, an electrochemical probe and
transmitter are illustrated for use with a bioreactor, in
accordance with an embodiment of the invention. The electrochemical
probe is connected to an electronics component that can be used to
interface the electrochemical probe with an existing (or
third-party) system or device configured to communicate with (or
take measurements from) a probe, such as, for example, a glass
probe. The pH probe includes a probe head at a distal end of a
probe body. The probe body is configured to extend into the
bioreactor for taking measurements of analytes of interests (e.g.,
H+ for pH measurements).
[0436] In some embodiments, a sensor is provided for insertion into
a container configured for use with a traditional or conventional
glass probe. In an embodiment, the container is cylindrical in
shape. In another embodiment, the container has a circular
cross-section. In another embodiment, the container is formed of
one or more metals. In another embodiment, the container is formed
of one or more metals including aluminum, nickel, platinum,
ruthenium, rhodium, tungsten, titanium, palladium, copper, silver,
gold and iron. In another embodiment, the container is formed of
one or more metals including stainless steel.
[0437] In an embodiment, an electrochemical sensor is provided
having a head portion comprising one or more working electrodes, a
counter electrode and a reference electrode, the head portion
mounted on a body portion, such as a tubular or cylindrical body
portion. The body portion can include threading configured to
permit the body portion to be mounted to a bioreactor. In an
embodiment, the threading includes PG-13 threading. In an
embodiment, the body portion can be formed of a polymeric material.
In another embodiment, the body portion can be formed of a metallic
material, such as stainless steel.
Calibration
[0438] Another aspect of the invention provides a system for
calibrating an electrochemical sensor with the aid of a robot. In
an embodiment, a robot is provided to obtain calibration
parameters, which can subsequently be provided to uses or stored in
an EEPROM onboard an electrochemical sensor.
[0439] In an embodiment, a robot comprises a robot arm for placing
an electrochemical sensor in a first solution or environment having
a known analyte, such as a solution or environment having a known
pH. The robot then measures the output of the electrochemical
sensor for the particular analyte measurements and records the
output, such as in a table. The robot then places the
electrochemical in a second solution or environment having a known
analyte, such as a solution or environment having a known pH. The
robot then measures the output of the electrochemical sensor for
the particular analyte measurement and records the output, such as
in a table. The robot can repeat this process, as desired, until a
predetermined number of measurements have been made. For example,
the robot can record electrochemical sensor outputs for pH 4, pH 7
and pH 10 solutions. The robot can then record the calibration data
in a calibration file for distribution to a user, or record the
calibration data in a memory chip, such as a memory chip onboard
the electrochemical sensor (e.g., EEPROM).
[0440] The pH sensors of the present invention are amenable to
miniaturization. In an embodiment, electrochemical sensors of the
present invention, such as pH sensors, can be miniaturized for use
in capsules for insertion in a subject's body. In another
embodiment, electrochemical sensors of the present invention, such
as pH sensors, can be miniaturized for use in capsules for
insertion in a subject's body. In another embodiment,
electrochemical sensors of the present invention, such as pH
sensors, can be miniaturized for use in skin patches applicable to
a subject's body. In another embodiment, electrochemical sensors of
the present invention, such as pH sensors, can be miniaturized for
use a nucleic acid sequencing array (e.g., 512.times.512
array).
Methods and Systems for Determining pH from Voltammetric or
Amperometric Data
[0441] Another aspect of the invention provides systems and methods
for converting peaks in voltammetric or amperometric data to pH
values. In some embodiments, a system is provided for implementing
the methods provided herein. Such methods can be implemented with
the aid of a system implementing an algorithm, as set forth in
machine-readable code.
[0442] In an embodiment, in a first step, the system identifies all
peaks having at least 10 fold greater intensity than background
noise level. A peak is defined as a current maximum that is
bordered by two current minimums, one minimum on each side of the
maximum. In an embodiment, a maximum can be 5% higher than a local
minimum. The system then ranks the maxima in terms of height (value
of the current at a given voltage), from maximum height to minimum
height. Next, using the largest maximum (i.e., the maximum having
largest value), the system takes the positions of the local minima
on each side of the largest maximum. The system then finds the
equation of the line that connects the two local minima. In an
embodiment, the line has the form of I=c+mV, wherein `I` is the
measured current, `V` is the applied voltage, `m` is the slope of
the line and `c` is the intercept. This process is repeated for
each data point in the amperometric data. For a set of three data
points a correction equation is provided. The system then subtracts
the correction equation for each original data point,
(I.sub.n,V.sub.n). A corrected data point is then achieved,
represented by (I.sub.nc, V.sub.n)=(I.sub.n-(c+m*V.sub.n),
V.sub.n). This correction removes other phenomena from the data,
such as local abnormalities due to noise.
[0443] Next, with a corrected data set, the system finds the
voltage of a maximum current of interest. In an embodiment, a
search for the voltage is limited to data points between two local
minima, as described above. The system first finds the difference
between a maximum and the minima. The system then takes the
difference between the higher minima and the maximum current. In an
embodiment, only the data that is above a predetermined percentage
of the current range of interest is used. In an embodiment, only
data that is above a 50%, or 55%, or 60%, or 65%, or 70%, or 75%,
or 80%, or 85%, or 90% of the current range of interest is used. In
another embodiment, only data that is from about 20% to 100% of the
current range of interest is used. For example, the upper 80% of
data between the higher minima and the maximum can be used. Next,
the data thus generated is fit to a parabola. While a parabola can
be computationally desirable, other mathematical functions can be
used to fit the data. For example, the data can be fit with a
Taylor Series expansion model to an order that minimizes the spread
among the data points and points generated by the model. Next, the
maximum of the fitted parabola (or other function) can be used as
the maximum of the amperometric curve. The distance between the
corrected maxima for the first working electrode (WE1) and the
second working electrode (WE2) is used to determine a pH. In an
embodiment, the pH can be represented by
pH=c+m(WE1-WE2)+m2(WE1-WE2).sup.2+m3(WE1-WE2).sup.3, wherein `c`,
`m`, `m2` and `m3` are constants that are determined
experimentally--i.e., the constants are determined by fitting the
equation to known one or more samples having known pH values).
Form Factors and Sensor Applications
[0444] Another aspect of the invention provides electrochemical
sensors having form factors for use with various applications, such
as insertion into a container for use with glass probe sensors.
This advantageously enables a user to replace glass probe sensor
with electrochemical sensors provided herein, such as redox-active
moiety containing sensors.
[0445] Sensors provided herein can be suited for various
applications. In some embodiments, sensors provided herein are
configured for use in bioreactors, such as, e.g., single-use and/or
disposable bioreactors. In other embodiments, sensors provided
herein are configured for use in sample preparation and/or
analytical systems, such as chromatography, including, for example,
liquid chromatography (e.g., high pressure liquid chromatography),
gas chromatography, affinity chromatography, supercritical fluid
chromatography, ion exchange chromatography, size-exclusion
chromatography, reversed phase chromatography, two-dimensional
chromatography, simulated moving-bed chromatography, pyrolysis gas
chromatography, fast protein liquid chromatography, countercurrent
chromatography, chiral chromatography, and column chromatography.
In other embodiments, sensors provided herein are configured for
use in process separation unit operations, such as distillation
columns and absorption columns. In other embodiments, sensors
provided herein are configured for use in continuous stirred tank
reactors. In other embodiments, sensors provided herein are
configured for use in plug flow rectors. In other embodiments,
sensors provided herein are configured for use in fluidized bed
reactors.
[0446] Sensors can have form factors (e.g., shapes, sizes) suited
for such various sensor applications provided herein. FIGS. 32A-32E
show exemplary sensors having form factors suited for various
applications. FIG. 32A shows a sensor having a form factor as found
in some conventional probes. A redox-active moiety containing
sensor 3201 is disposed at a probe tip of the sensor of FIG. 32A.
FIG. 32B shows an electrochemical sensor having a form factor that
is suited for use in pH sensors. The sensor of FIG. 32B can be used
at the probe tip of the sensor of FIG. 32A. FIG. 32C shows a
substantially flat sensor. The sensor of FIG. 32C can be suited for
use in reactors, such as bioreactors. The sensor of FIG. 32C can
have a distribution of working electrodes as described herein. FIG.
32D shows a sensor configured for use with in-line flow systems,
such as, for example, plug flow reactors. A redox-active moiety
containing sensor 3202 is disposed at a probe tip of the sensor of
FIG. 32D. FIG. 32E shows a sensor configured for use in a system
having a predetermined volume, such as, e.g., about 30 microliters
of retained volume.
[0447] Sensors provided herein can have supports for enabling the
sensors to be used in various applications. For example, the
sensors of FIGS. 32A, 32D and 32E can have threads, o-rings, and
hex bodies that allow for the sensors to be mounted in various
settings, such as, for example, reactors (e.g., bioreactors),
various unit operations (e.g., distillation columns), flow-through
systems and fermentors.
EXAMPLES
Reagents and Instrumentations
[0448] Vinylferrocene, vinylanthracene, hydrofluoric acid were
purchased from Sigma-Aldrich (Sigma-Aldrich INC., USA),
ferrocenecarboxaldehyde and mesitylene were obtained from Alfa
Aesar (Alfa Aesar INC., USA), and 9-anthracene-carboxaldehyde was
obtained from Acros Organics (Acros Organics INC. USA). All the
chemicals were obtained with the highest grade available and were
used without further purification.
[0449] Different single-side polished, primary flat, 500 .mu.m
thick silicon wafers with (111) and (100) orientation were
purchased from Virginia Semiconductor with the following
specification: i) P-type (100, 10-90 .OMEGA.-cm resistivity), ii)
P-type (100, 0.001-0.005 .OMEGA.-cm resistivity), iii) N-type (100,
10-40 .OMEGA.-cm resistivity), iv) N-type (100, 0.02-0.05
.OMEGA.-cm resistivity), v) P-type (111, 0.001-0.004 .OMEGA.-cm
resistivity) and yl) N-type (111, 0.001-0.005 .OMEGA.-cm
resistivity).
[0450] Electrochemical measurements were recorded using an Autolab
computer controlled potentiostat (Ecochemie, Utrecht, Netherlands)
with a standard three-electrode configuration, consisting of a
saturated calomel reference electrode (SCE, Radiometer, Copenhagen,
Denmark), a platinum auxiliary electrode (Bioanalytical Systems
INC., USA)) and silicon (Virginia Semiconductor INC, USA) working
electrode.
[0451] Different pH solutions in the range of 1 to 12 were also
prepared in deionized water as follows: pH 1.2, 0.10 M perchloric
acid; pH 2.2, 0.05 M perchloric acid; pH 4.6, 0.1 M acetic
acid+0.10 M sodium acetate; pH 5.6, 0.5 M sodium acetate; pH 6.5,
0.025 M K.sub.2PO.sub.4+0.025 M KH.sub.2PO.sub.4; pH 7.33, 0.05 M
K.sub.2PO.sub.4; pH 9.3, 0.10 M sodium borate; pH 13.5, 0.1 M
sodium hydroxide. These solutions also contained an addition of
0.10 M sodium perchlorate as supporting electrolyte. The pH of
these solutions was measured using the SevinMulti (Mettler Toledo)
pH meter.
Example 1
Preparation of H-Terminated Silicon Surface
[0452] Silicon wafers (oriented (111) or (100), cut into ca.
1.times.1 cm.sup.2 pieces) were cleaned using "Piranha" solution
(concentrated H.sub.2SO.sub.4:30% H.sub.2O.sub.2, 3:1, v/v) at
about 80.degree. C. for 30 min and rinsed thoroughly with deionized
water. (In some cases, smaller pieces, e.g., 2 mm.times.3.3 mm or 2
mm.times.2.7 mm were used.) Subsequently, the wafer pieces were
oxidized in H.sub.2O.sub.2:HCl:H.sub.2O (2:1:8) at about 80.degree.
C. for 15 min, and in H.sub.2O.sub.2:NH.sub.4OH:H.sub.2O (2:1:8) at
about 80.degree. C. for another 15 min, rinsed copiously with
deionized water. The cleaned Si(100) wafers pieces were then etched
in 2.5% HF solution for about 15 min. These procedures eliminate
the native silicon oxide layer, yielding an H-terminated surface.
The H-terminated substrates were quickly rinsed with deionized
water, dried with nitrogen gas and were used immediately for the
derivatization experiments. The Si(100) (10-90 .OMEGA.-cm, P-type)
was used for the experiments below.
Example 2
Derivatization of H-Terminated Silicon Surface with Ferrocene
Moieties
[0453] Approximately 10 mmol mesitylene solution of vinylferrocene
(VFc) or ferrocenecarboxaldehyde (FcA) was put in a round bottom
flask and bubbled with nitrogen or argon gas for at least 30 min. A
piece of the H-terminated silicon substrate was then immersed in
the solution and allowed to react with VFc or FcA for about 12 h
under reflux at about 150.degree. C. in an oil bath. During the
reaction, the solution was also purged with nitrogen (or argon) to
eliminate dissolved oxygen and to prevent the substrate from being
oxidized. After the reaction, the substrate derivatized with VFc or
FcA was rinsed with dichloromethane, acetonitrile, and methanol;
and dried under a stream of nitrogen gas. The derivatization of the
H-terminated surface with ferrocene moieties as described in
Example 2 is illustrated in FIG. 5.
Example 3
Derivatization of H-Terminated Silicon Surface with Anthracene
Moieties
[0454] Approximately 10 mmol mesitylene solution of vinylanthracene
(VA) or anthraldehyde (AnA) was put in a round bottom flask and
bubbled with nitrogen or argon gas for at least 30 min. A piece of
the H-terminated silicon substrate was then immersed in the
solution and allowed to react with VA or AnA for about 12 h under
reflux at about 150.degree. C. in an oil bath. During the reaction,
the solution was also purged with nitrogen (or argon) to eliminate
dissolved oxygen and to prevent the substrate from being oxidized.
After the reaction, the substrate derivatized with VA or AnA was
rinsed with dichloromethane, acetonitrile and methanol; and dried
under a stream of nitrogen gas. The derivatization of the
H-terminated surface with anthracene moieties as described in
Example 3 is illustrated in FIG. 6.
Example 4
Derivatization of H-Terminated Silicon Surface with Both the
Anthracene and Ferrocene Moieties
[0455] A 10 mmol mesitylene solution of anthracene (VA or AnA) and
ferrocene (VFc or FcA) in 1:1 ratio was put in a round bottom flask
and bubbled with nitrogen or argon gas for at least 30 min. A piece
of the H-terminated silicon substrate was then immersed in the
solution and allowed to react with the anthracene and ferrocene
mixtures for about 12 hours under reflux at 150.degree. C. in oil
bath. During the reaction, the solution was also purged with
nitrogen (or argon) to eliminate dissolved oxygen and to prevent
the substrate from being oxidized. After the reaction, the
derivatized substrate was rinsed with dichloromethane, acetonitrile
and methanol, and dried under a stream of nitrogen gas. FIG. 7
illustrates a reaction in which the silicon surface derivatized
with both the anthracene and ferrocene using VFc and VA.
Example 5
Electrochemical Measurements of the Derivatized Silicon Wafers in
Different pH Solutions
[0456] Square wave voltammetry (SWV) was carried out for the
derivatized silicon wafers in a specially designed electrochemical
cell as shown in FIG. 8. The electrochemical measurements were
performed using a standard three-electrode configuration. In these
experiments, the derivatized silicon wafers were used as the
working electrode, and was exposed to different pH solutions (about
10 mL) in the electrochemical cell. SWV were performed with a
frequency of 10 Hz, a step potential of 2 mV and an amplitude of 25
mV.
[0457] The amperometric response of the anthracene or ferrocene
derivatized silicon substrate at different pH solutions was studied
using SWV. SWV was used as the electrochemical probe of the system
because it produces a well-defined voltammetric peak in a single
sweep. The corresponding square wave voltammograms recorded using a
derivatized ferrocene electrode at different pH solutions from pH
1.23 to 9.33 are shown in FIG. 9. These voltammograms show that as
the pH values increase, the peak potential of the ferrocene peaks
remain at the same peak potential. These results show that
ferrocene is a pH insensitive molecule which can act as an internal
reference material.
[0458] The corresponding SW voltammograms recorded using a
derivatized anthracene electrode at different pH solutions, from pH
1.23 to 13.63, are shown in FIG. 10 (a). These voltammograms show
that as the pH value increases, the peak potential attributed to
the anthracene shifts to a more negative potential. The
corresponding plot of the peak potential against different pH is
given in FIG. 10(b). The plot reveals a linear response from pH 1
to pH 14 with a corresponding gradient of ca 55.1 mV per pH unit.
The ability of anthracene compounds to act as pH sensitive molecule
is thus demonstrated.
[0459] The corresponding SW voltammograms recorded using a
derivatized ferrocene+anthracene electrode at different pH
solutions, from pH 1.23 to 9.33, are shown in FIG. 11 (a). These
voltammograms illustrate that as the pH values increase, the
ferrocene peak remains at the same peak potential while the
anthracene peak shifts to a more negative potential. The
corresponding plot of the difference between the two peak
potentials versus pH is shown in FIG. 11(b). The plot reveals a
linear response from pH 1 to pH 9.33 with a corresponding gradient
of ca 45.1 mV per pH unit.
Example 6
Heat Stability
[0460] A SW voltammogram was recorded for the silicon wafer
derivatized with both VA and FcA moieties at room temperature in pH
6.52 buffer. The derivatized silicon sample was then autoclaved in
a Consolidated Stills & Sterilizers for 40 min, and a SW
voltammogram was recorded in pH 6.52 buffer after the autoclave.
Next, the same sample was autoclaved again under the same condition
10 times, with a SW voltammogram recorded in pH 6.52 buffer after
each autoclave.
[0461] The heat stability testing of the pH sensor was performed
using a FcA+VA derivatized silicon sample. The resultant
voltammograms are shown in FIG. 12. A decrease in both the
ferrocene and anthracene currents were observed after the first
autoclave. Thereafter, both the peak currents remain relatively
stable though subsequent cycles of autoclaving; in fact the peaks
remain stable for ten cycles, showing that the sensor can withstand
repeated heat sterilization.
Example 7
Fouling Test
[0462] Four SW voltammograms were recorded for the four separate
silicon wafers derivatized with FcA and AnA moieties at room
temperature in pH 6.52 buffer. These derivatized samples were then
autoclaved in the Consolidated Stills & Sterilizers for 20 min
and were immersed in a 5 mL cell culture fermentation medium for
six days. Then these samples were taken out of the cell culture
medium, and SW voltammograms were recorded in pH 6.52 buffer again.
SW voltammetry was also carried out in the cell culture medium,
using the same silicon wafers.
[0463] The fouling testing of the pH sensor was performed using
five derivatized silicon samples. The resultant voltammograms were
shown in FIG. 13. In all cases, the anthracene peaks remain stable
with after six days exposure. The ferrocene peaks decrease after
exposure, but the ferrocene peaks are still identifiable. These
findings demonstrate that the pH sensors are still in good working
order after six days exposure to the cell culture environment,
demonstrating that the ability of the derivatized sensor to resist
fouling. A control sensor was incubated for 6 days in culture fluid
without cells or secreted proteins. This sensor's voltammogram
exhibited a similar profile to the four that had been incubated in
the actual cell fermentation environment, (FIG. 14) suggesting that
any loss of signal amplitude was not a function of cellular debris
deposition.
Example 8
Stability of the Anthracene- and Ferrocene-Derivatized Si
Surfaces
[0464] The stability testing of the Ac+Fc derivatized silicon
surface was conducted for 22 days under continuous electrochemical
measurement using PGSTAT12 autolab potentiostat in pH 4.65 buffer
medium, scanning from -0.8 V to 0.8 V vs. Ag. The Ac peak remained
as a well-defined peak with a full width half maximum (FWHM) of
.about.60 mV throughout the course of the experiments, while the Fc
peaks broadened with time. Although the Fc peaks became broader,
they are still identifiable and can be used as a reference. These
findings demonstrate that the two-component derivatized silicon
surface are still in good working order after 22 days continuous
operation in pH 4.65 buffer medium, demonstrating the long term
stability of the derivatized surface in the buffer medium.
[0465] In general, the Fc peaks are most well-defined and stable
when the Fc molecules were derivatized onto a heavily doped silicon
substrate. FIG. 15(a) depicts SW voltammetric responses FcA on
Si(100, N-type, 1-5 m.OMEGA. cm) in pH 7.33 buffer medium, showing
every 50.sup.th scan of the 2,500 consecutive runs.; FIG. 15(b)
depicts voltammetric responses of VFc on Si(111, N-type, 0.02-0.05
cm in pH 7.33 buffer medium, showing every 50.sup.th scan of the
2,500 consecutive runs. In some cases, both VFc and FcA moieties
behave better on an N-type substrate than a P-type substrate in
terms of the size of the peak current produced. The Fc derivatized
silicon surfaces were all pH insensitive, i.e., the Fc peak does
not shift upon exposure to different pH environments.
Example 9
Temperature Variation
[0466] The Nernst equation provides a theoretical framework for
evaluating the temperature dependence of redox active species. It
predicts that the slope of the peak potential against pH plot will
increase as the temperature increases:
E p = E f 0 - 2.3 RTm nF pH Eq . 1 ##EQU00001##
[0467] Where E.sub.p is the peak potential (V), E.sub.f.sup.0 is
the standard electrode potential (V), R is the universal gas
constant (J K.sup.-1mol.sup.-1), T is the absolute temperature (K),
F is the Faraday constant (C mol.sup.-1), m and n are the number of
protons and electrons involved in the redox reaction, respectively.
In the case of anthracene, the redox process for such molecules in
the aqueous solution involves the participation of 2 electrons and
2 protons, thus m=n=2.
[0468] SW voltammograms were recorded for three pH solutions at pH
4.65, 7.33 and 9.35. FIG. 16 shows the overlaid SW voltammograms of
Ac derivatized silicon over the temperature range of 8 to
56.degree. C. in pH 7.33 buffer medium. Similar responses were
obtained at pH 4.65 and pH 9.35. There is a shift of the peak
potential to more negative values with increasing temperature which
may be attributed in part to a combination of changes in the
reference couple, the temperature dependence of the formal
potential (E.sub.f.sup.0), and the temperature term in Eq. 1.
Analysis of the slope of the peak potential as a function of pH
obtained experimentally at each temperature is tabulated in Table
1, which illustrates that the slope of pH against peak potential
plot varies with temperature. The theoretical slope as predicted by
the Nernst equation as seen in Eq. 1 is also listed in Table 1 for
comparison. As can be seen, the variation of the gradient of peak
potential with pH is not Nernstian and indeed is relatively
insensitive of temperature varying by about 2 mV/pH unit over a
temperature range of .about.50.degree. C. This dependence can be
compared to a 10 mV/pH shift that can be observed for a glass
electrode. This small shift with temperature is beneficial in that
it not only demonstrates that these Ac derivatized silicon wafers
may be used as pH sensors at elevated temperatures, but also that
they are not greatly affected by changes in temperature.
TABLE-US-00001 TABLE 1 A comparison of the theoretically predicted
slope and experimentally obtained slope of the plot of pH against
Ac peak potential for a range of temperatures. T (.degree. C.) 8 17
28 44 56 T (K) 281 290 301 317 329 Theoretical (mV/pH) 55.7 57.5
59.7 62.8 65.2 Experimental 55.0 55.5 56 56.3 57.0 (mV/pH)
Example 10
Stability During Active Measurement in Cell Culture Medium
[0469] A fouling testing was carried out using anthracene
derivatized silicon (100) wafer in the incubator (under controlled
temperature of 37.degree. C. and CO.sub.2 content of 5%) with
continuous electrochemical measurements in cell culture (LP VA)
medium for 7 days using a three-electrode setup connected to an
.mu.autolab type III potentiostat. The three-electrode setup (in an
electrochemical cell) was autoclaved in a Consolidated Stills &
Sterilizers autoclave for 40 min prior to adding 5 mL of the cell
culture medium to the autoclaved setup. The electrochemical cell
containing the medium was then transferred to the incubator, where
continuous electrochemical measurements were performed.
[0470] Voltammograms were taken repeatedly over 7 days resulting in
10,000 consecutive voltammograms. FIG. 18(a) shows voltammograms
taken over the 7 day period (every 250.sup.th scan of the 10,000
consecutive runs). The anthracene peak was observed at
.about.-0.71V vs. Ag and remained consistent throughout the 7 days
of in-situ electrochemical measurements in the cell culture medium.
A plot of the anthracene peak potential over the 7 day time period
is shown in FIG. 18(b). These findings demonstrate that the
anthracene derivatized silicon is still in good working order and
that the peak potential remains consistent while the sensor is
actively sensing the solution for 7 days.
Example 11
Derivatization of H-Terminated Silicon Surfaces with
2-Allyl-1-Hydroxy-Anthraquinone
[0471] Into a round bottom flask containing 50 mL of mesitylene was
placed 5 mg of 2-allyl-1-hydroxy-anthraquinone. Nitrogen was
bubbled for at least 30 minutes. Subsequently, a H-terminated
silicon wafer was placed into the flask and was allowed to react
with 2-allyl-1-hydroxy-anthraquinone for 12 hours under reflux at
150.degree. C. in an oil bath. During the reaction, the solution
was purged with nitrogen gas to prevent the substrate from being
re-oxidized. Following the reaction, the
2-allyl-1-hydroxy-anthraquinone derivatized silicon wafer was
rinsed with dichloromethane, hexane, then methanol and dried under
a stream of nitrogen.
Example 12
Derivatization of Silicon Wafers with Ferrocene Using a
Semiconductor Oxide Surface
[0472] Si(100) wafer pieces were cleaned using "Piranha" solution
(3:1 v/v concentrated H2SO4/30% H2O2) for 30 minutes at 100.degree.
C. and rinsed thoroughly with deionized water. Subsequently, the
wafers were oxidized in 2:1:8 HCl/H2O2/H2O for 15 minutes at
80.degree. C. followed by 2:1:8 NH4OH/H2O2/H2O for 15 minutes at
80.degree. C. and then rinsed with copious amounts of water. The
oxidized silicon wafers were then immersed in 2%
3-aminopropyltriethoxysilane (APTES) solution in acetone for 2
minutes followed by rinsing with acetone to form an
amino-terminated monolayer. This was then allowed to react with 10
mg/mL dicyclohexyl-carbodiimide (DCC) and 50 mg/mL ferrocene
carboxylic acid in DMSO for 12 hours. Following the reaction, the
ferrocene derivatized silicon wafer was rinsed with
dichloromethane, acetone, and methanol and then dried under a
stream of nitrogen.
Example 13
Derivatization of Silicon Wafers with Anthracene Using a
Semiconductor Oxide Surface
[0473] Si(100) wafer pieces were cleaned using "Piranha" solution
(3:1 v/v concentrated H2SO4/30% H2O2) for 30 minutes at 100.degree.
C. and rinsed thoroughly with deionized water. Subsequently, the
wafers were oxidized in 2:1:8 HCl/H2O2/H2O for 15 minutes at
80.degree. C. followed by 2:1:8 NH4OH/H2O2/H2O for 15 minutes at
80.degree. C. and then rinsed with copious amounts of water. The
oxidized silicon wafers were then immersed in 2% 3-APTES solution
in acetone for 2 minutes followed by rinsing with acetone to form
an amino-terminated monolayer. This was then allowed to react with
10 mg/mL dicyclohexyl-carbodiimide (DCC) and 50 mg/mL anthracene
carboxylic acid in DMSO for 12 hours. Following the reaction, the
anthracene derivatized silicon wafer was rinsed with
dichloromethane, acetone and methanol and then dried under a stream
of nitrogen.
Example 14
Ferrocene Covalently Attached to Various Doped Si(100)
Substrates
[0474] Ferrocene was covalently attached to the following four
silicon surfaces via an H-terminated silicon substrate and vinyl
ferrocene: i) Si (100) N-type (0.02-0.05 ohm-cm)--highly-doped
N-type, ii) Si (100) P-type (0.005-0.020 ohm-cm)--highly-doped
P-type, iii) Si (100) N-type (10-40 ohm-cm)--lightly-doped N-type,
and iv) Si (100) P-type (10-90 ohm-cm)--highly-doped P-type. A
well-defined ferrocene peak was observed for all four substrates.
The highly-doped substrates in general produced a larger
electrochemical current than the corresponding lightly-doped
substrates. While not being bound by theory, this difference may be
explained by the fact that the highly-doped substrates contain more
charge carriers, i.e., are more conductive. Similar observations
were observed when the electrochemistry was performed in a solution
of ferrocene carboxaldehyde. FIG. 19 has charts showing the peak
current of silicon substrates derivatized with (a) vinyl ferrocene
and (b) ferrocene carboxaldehyde in pH 1.63 solution on the four
types of doped silicon.
Example 15
Four Electrode System
[0475] A system was constructed having four electrodes comprising
(i) a counter electrode, (ii) a reference electrode, (iii) a
highly-doped N-type silicon wafer derivatized with vinyl ferrocene,
and (iv) a lightly-doped P-type silicon wafer derivatized with
anthracene carboxaldehyde. FIG. 20 illustrates the voltammograms
obtained with the four electrode system consisting of a
highly-doped N-type silicon wafer derivatized with vinyl ferrocene
and a lightly-doped P-type silicon wafer derivatized with
anthracene carboxaldehyde. The voltammograms were obtained in pH
7.03 solution and the peak potentials were stable over 200
consecutive scans (every 50th scan is shown).
Example 16
Working Electrodes Coated with Nafion Membranes
[0476] A Nafion membrane was applied to a working electrode having
ferrocene. A 5 mil thick section of pre-formed Nafion membrane
(N115) was pre-treated either by soaking in water or in an acid
solution (about 5% HCl or 5M HNO.sub.3) at about 100.degree. C. for
about 1 hr. The treated membrane was then cut to a size appropriate
to cover the surface of the working electrode. The Nafion membrane
was then coated with a liquid dispersion of Nafion 117 or Nafion
2020, and applied directly to the electrode surface. The
membrane-coated electrode was then cured in a regular or vacuum
oven at about 120.degree. C. for about 1 hr. The electrode was then
rehydrated and tested. Testing showed stability as to the peak
position in SWV.
Example 17
Nafion-Plastic Composites
[0477] A porous plastic membrane (pore size 75-100 micron, 0.5 mm
thick) is cut into a 1 cm disk. The disk is saturated with a Nafion
dispersion and allowed to dry for 30 min at 50.degree. C. After a
second cycle of saturation and drying, the disk is placed in a
120.degree. C. oven for about 1 hour. A single electrode is then
wetted with the Nafion dispersion and placed on the smooth side of
the disk. This assembly is then cured at 120.degree. C. for about 1
hour. The backside of the electrode is then coated with a
conductive epoxy and a wire is affixed. The assembly is potted into
a waterproof fixture for electrochemical measurements.
Example 18
Light Sensitivity
[0478] A sensor has a first working electrode (WE1) and second
working electrode (WE2) that are formed of silicon. WE1 includes a
layer of ferrocene and WE2 includes a layer of anthracene. A first
probe has WE1 and WE2 that are shielded from light with PES layers,
and a second probe has WE1 and WE2 that are not shielded (or
unshielded) from light. The first probe and second probe are
inserted into a solution having a pH of about 4. Continuous pH
measurements are made and plotted as the difference between the
peak potentials of WE1 and WE2. Sensor outputs (y-axis, mV) as a
function of time .alpha.-axis) during pH measurements for both the
shielded (dashed line, top) and unshielded (solid line, bottom)
probes are shown in FIG. 30. The output of the shielded sensor upon
exposure to light is different than the output of the unshielded
sensor upon exposure to light. During pH measurements, the output
of the unshielded sensor shows a sensor response that is coincident
with exposure to light; the output of the shielded sensor does not
exhibit such a behavior. Sensor output when the first and second
working electrodes are shielded (top) does not display the
light-sensitive behavior of the unshielded working electrodes
(bottom). The figure also shows the effect of either infrared (IR)
or room light for both shielded and unshielded probes. The shielded
sensor is unresponsive to IR or room light.
Example 19
The Effect of Light on Sensor Output
[0479] A sensor has a first working electrode (WE1) and second
working electrode (WE2) that are formed of silicon. WE1 includes a
layer of ferrocene and WE2 includes a layer of anthracene. The
sensor is immersed in various solutions, each solution having a
predetermined pH. Sensor output is recorded both with and without
exposure of the sensor to light. FIGS. 31A-31F show sensor output
(y-axis; current, arbitrary units) at various pH's and under light
and dark conditions as a function of voltage (mV). Channel 2
corresponds to the sensor output for WE2. Also provided in the
figures are Channel 2 peak positions (mV). FIG. 31A shows the
sensor output at pH 5 without exposure to light (i.e., the sensor
is in the dark). FIG. 31B shows the sensor output at pH 5 while the
sensor is exposed to light. FIG. 31C shows the sensor output at pH
7 without exposure to light. FIG. 31D shows the sensor output at pH
7 while the sensor is exposed to light. FIG. 31E shows the sensor
output at pH 10 without exposure to light. FIG. 31F shows the
sensor output at pH 10 while the sensor is exposed to light. The
Channel 2 signal (current) when the sensor is exposed to light is
more intense than situations in which pH measurements are made
without exposure to light. This indicates that, when the sensor is
exposed to light, better signal to noise may be achieved.
Example 20
The Effect of Light on Sensor Output
[0480] A sensor has a first working electrode (WE1) and second
working electrode (WE2) that are formed of silicon. WE1 includes a
layer of ferrocene and WE2 includes a layer of anthracene. The
sensor is immersed in a solution with a pH of 5. Sensor output is
recorded both with and without exposure of the sensor (and WE1 and
WE2) to light of fixed intensities and wavelengths. Light is
exposed on sensor surfaces having the ferrocene and anthracene
moieties. It is observed that peak shapes are sharper in the
presence of light up to a certain intensity and wavelength. At
higher intensities the signal is destroyed. Optimal responses are
obtained in the IR or near-IR wavelength range, such as light
having a wavelength greater than or equal to about 750 nm, or
greater than or equal to about 850 nm.
Example 21
Doping Configurations
[0481] Multiple sensors are formed from silicon wafers having
varying doping configurations. Each sensor has a first working
electrode (WE1) and second working electrode (WE2) that are formed
of silicon. WE 1 includes a layer of ferrocene and WE2 includes a
layer of anthracene. During pH measurements, electrochemical
responses from anthracene-covered electrodes (WE2) are observed
when anthracene is derivatized onto the following silicon surfaces:
Si(100) wafer, p-type, resistivity between about 1 and 20
.OMEGA.-cm; Si(100) wafer, p-type, resistivity between about 1 and
90 .OMEGA.-cm; Si(111) wafer, p-type, resistivity between about 1
and 20 .OMEGA.-cm; and Si(111) wafer, p-type, resistivity between
about 1 and 10 .OMEGA.-cm. An anthracene signal with an appreciable
signal-to-noise ratio is not observed when derivatized on an n-type
silicon substrate. An anthracene signal is not observed when
derivatized on a p-type silicon substrate with resistivity less
than about 1 .OMEGA.-cm. Electrochemical responses are observed for
ferrocene on both p-type and n-type silicon (both Si(100) and
Si(111). Signals for Ferrocene on all resistivities of p-type
silicon and on n-type silicon with resistivities <5 .OMEGA.-cm
are unstable over time (in some situations, it is observed that the
signal degrades with continuous square wave voltammetric scanning)
Ferrocene on n-type surfaces with resistivities between 1 and 90
.OMEGA.-cm are stable.
Example 22
Sensor Stability
[0482] A test was conducted to monitor fermentation in a
fermentation reactor. The pH of the reactant (i.e., cell culture)
and product content of the fermentation reactor was monitored over
a 10-day period using both a redox-active moiety-based pH sensor
(see above) and a conventional glass electrode sensor (Applikon
Biotechnology). Both sensors were autoclaved within the bioreactor
prior to the initiation of the cell culture. The redox-active
moiety-based pH sensor was not recalibrated during the 10-day
period. FIG. 33 shows the pH of the contents of the fermentation
reactor over the 10-day period. The redox-active moiety-based pH
sensor tracked the glass electrode sensor without any appreciable
drift and without exhibiting any appreciable electronic background
noise throughout the course of the 10-day period. In contrast, the
glass electrode sensor exhibited electronic noise during the 10-day
period.
[0483] Electrochemical sensor systems, devices and methods provided
herein can be combined with or modified by other systems, devices
and methods. For example, electrochemical sensors described herein,
including methods for forming such sensors, can be combined with or
modified by systems and methods described in U.S. patent
application Ser. No. 12/049,230 to Kahn et al. ("SILICON
ELECTROCHEMICAL SENSORS") and PCT/US2008/066165 to Kahn et al.
("SEMICONDUCTOR ELECTROCHEMICAL SENSORS"), which applications are
entirely incorporated herein by reference.
[0484] In some embodiments, sensors have been described as being
used to detect the presence or absence of an analyte (e.g., H+). It
will be appreciated, however, that detecting the presence or
absence of an analyte can include detecting (or measuring) the
concentration of an analyte. For example, detecting the presence or
absence of H+ in a liquid sample using any of the sensors described
herein can include detecting (or measuring) the concentration of
H+.
[0485] It should be understood from the foregoing that, while
particular implementations have been illustrated and described,
various modifications can be made thereto and are contemplated
herein. It is also not intended that the invention be limited by
the specific examples provided within the specification. While the
invention has been described with reference to the aforementioned
specification, the descriptions and illustrations of the preferable
embodiments herein are not meant to be construed in a limiting
sense. Furthermore, it shall be understood that all aspects of the
invention are not limited to the specific depictions,
configurations or relative proportions set forth herein which
depend upon a variety of conditions and variables. Various
modifications in form and detail of the embodiments of the
invention will be apparent to a person skilled in the art. It is
therefore contemplated that the invention shall also cover any such
modifications, variations and equivalents.
* * * * *