U.S. patent application number 11/054564 was filed with the patent office on 2006-08-10 for optoelectronic probe.
Invention is credited to Haian Lin.
Application Number | 20060175192 11/054564 |
Document ID | / |
Family ID | 36758588 |
Filed Date | 2006-08-10 |
United States Patent
Application |
20060175192 |
Kind Code |
A1 |
Lin; Haian |
August 10, 2006 |
OPTOELECTRONIC PROBE
Abstract
The present invention, referred to as optoelectronic probe,
concerns a novel apparatus and method for characterization and
micromanipulation of particles or biomolecules in an electrolyte
solution. Electric fields, which include both time constant and
time-varying components, are applied to a thin insulating layer
covered, lightly doped semiconductor material. Illumination injects
carriers into the insulator/semiconductor interface to compensate
the leaking minority carrier current and maintain an inversion
layer, which works as an electrode to control the particle
movements. A particle array, or even a single cell, can be
assembled in, or moved along with the inversion layer electrode,
which is induced by illumination. Furthermore, an impedance
analyzer is utilized to characterize the trapped particles, or
single cell. The present invention has numerous uses, such as
bio-chemical analysis systems, and nanosize structures assembly for
electronic or optical devices.
Inventors: |
Lin; Haian; (Bethelehem,
PA) |
Correspondence
Address: |
HAIAN LIN
5019 PREAKNESS PLACE
BETHELEHEM
PA
18020
US
|
Family ID: |
36758588 |
Appl. No.: |
11/054564 |
Filed: |
February 9, 2005 |
Current U.S.
Class: |
204/194 ;
204/415; 324/71.1; 324/754.23 |
Current CPC
Class: |
G01R 1/071 20130101 |
Class at
Publication: |
204/194 ;
204/415; 324/753; 324/071.1 |
International
Class: |
C25D 1/00 20060101
C25D001/00 |
Claims
1. A method for controlling the movement of particles in an
electrolyte, comprising the following steps: providing a
semiconductor material; forming an insulating layer over said
semiconductor material; attaching an electrolyte solution to said
insulating layer; applying an electric input to said semiconductor
material, said electric input having a predetermined polarity;
illuminating the surface of said semiconductor material with a
predetermined light beam; maintaining an inversion region at the
interface between said semiconductor material and said insulating
layer, said inversion region being defined by said predetermined
light beam, said inversion region working as an electrode; and
providing a plurality of particles suspended in said electrolyte
solution, said particles being manipulated by said electrode, in
accordance with said electric input and said predetermined light
beam.
2. The method of claim 1, further comprising a characterizing step
which is performed using an instrument to measure the electric
response of said particles on said electric input.
3. The method of claim 2, wherein said instrument is an impedance
analyzer.
4. The method of claim 1, wherein said semiconductor material has a
predetermined doping density to form a depletion layer at the
interface between said semiconductor material and said insulating
layer.
5. The method of claim 1, wherein said insulating layer has a
predetermined thickness to enable to tunnel carriers between said
electrolyte and said semiconductor material.
6. The method of claim 1, wherein said electric input is a voltage
input that comprises both a time constant component and a
time-varying component.
7. The method of claim 6, wherein said time-varying component has a
frequency to polarize said particles.
8. The method of claim 7, further comprising the step of sweeping
said frequency from a first predetermined value to a second
predetermined value.
9. The method of claim 6, wherein said time-varying component is a
direct current time-varying component which has the same polarity
as said time constant component.
10. The method of claim 1, further comprising the step of
assembling an array of said particles over said electrode, in
accordance with said electric input and said predetermined light
beam.
11. The method of claim 10, further comprising the step of
maintaining said array by either maintaining said predetermined
light beam and said electric input, chemically linking said
particles, or confining said particles.
12. The method of claim 10, further comprising the step of
disassembling said arrays by removing said electric input, or
turning off said predetermined light beam.
13. The method of claim 10, further comprising the step of moving
said array by steering said predetermined light beam.
14. The method of claim 10, wherein said predetermined light beam
is adjusted to reconfigure said particle array in accordance with
said predetermined light beam.
15. A method for probing particles in an electrolyte using a
semiconductor material covered with an insulating layer,
comprising: applying an electric input to said semiconductor
material; illuminating the surface of said semiconductor material
with a predetermined light beam to form an inversion region at the
interface between said insulating layer and said semiconductor,
said inversion region working as an electrode; trapping a particle
over said electrode in accordance with said electric input; and
analyzing the electric response of said particle on said electric
input.
16. A method for characterizing particles in an electrolyte using
photosensitive material, comprising: applying an electric input on
said photosensitive material; illuminating the surface of said
photosensitive material with a predetermined light beam in
conjunction with said electrical input to generate an electrode to
control the movement of said particles; and analyzing the electric
response of said particles on said electric input.
17. An apparatus for probing particles in an electrolyte,
comprising: a semiconductor material; an insulating layer formed
over said semiconductor material; an electrolyte solution attached
to said insulating layer; an electric source which applies an
electric input to said semiconductor material, said electric input
having a predetermined polarity; a predetermined light beam which
illuminates the surface of said semiconductor material to maintain
an inversion region at the interface between said insulating layer
and said semiconductor, said inversion region working as an
electrode; a plurality of particles suspended in said electrolyte
solution, said particles being manipulated by said electrode in
accordance with said electric input; and an instrument for
measuring the electric response of said particles on said electric
input.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] Not Applicable
STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT
[0002] Not Applicable
REFERENCE TO SEQUENCE LISTING, A TABLE, OR A COMPUTER PROGRAM
LISTING COMPACT DISK APPENDIX
[0003] Not Applicable
BACKGROUND OF THE INVENTION
Field of Invention
[0004] The present invention is directed generally to methods and
apparatus based on optoelectronic effect,
electro/dielectro-phoresis, and impedance spectroscopy, in order to
trap, move, deform and characterize particles, such as cells,
molecules, any type of colloids, any of inorganic and bio-organic
substances, beads, as well as pucks and like small things.
"Optoelectronic Probe" refers to the invention described
herein.
BACKGROUND OF THE INVENTION
[0005] I. Optical Tweezers
[0006] The manipulation of micro- or nano-size particles is
considered as the key for the new generation of photonic,
optoelectronic, and electronic devices, as well as biochemical
analysis systems. Optical tweezers is one of the most unique
invention in this area and was first successfully demonstrated by
A. Ashkin et al. in pioneering works in 1985. (Ashkin, A.;
Dziedzic, J. M., "Observation of Radiation-Pressure Trapping of
Particles by Alternating Light Beams", Phys. Rev. Lett. 54, pp
1245-1248 (1985)) The technique of optical tweezers is based on the
forces of radiation pressure. These are dipole- or gradient-forces
arising from the momentum of the light itself. To make these forces
large enough to accelerate, decelerate, deflect, guide, and even
stably trap small particles, one has to use continuous wave
coherent laser beams to achieve the high intensities and high
intensity gradients. Combined with other techniques, optical
tweezers can also be a unique tool to characterize the trapped
particle. For example, laser fluorescence techniques give increased
opportunities to a proper identification of different types of
biological objects or labeling.
[0007] Although the Optical Tweezers is a very powerful tool, it
also has its limitations, such as: 1) that the trapping zone is
rather small (on the order of the light wavelength); and 2)
focusing the beam leads to very high intensities that can endanger
the integrity of biological objects.
[0008] II. Electrophoresis/Dielectrophoresis Based Arts
[0009] When it is exposed to an electrical field, a charged
particle will experience a force and the resulting motion is called
as electrophoresis (EP). A neutral particle can also be polarized
under electrical field. If a nonuniform direct current (DC) or
alternating current (AC) field exists, the polarized particle will
move towards or away from regions of high electric-field intensity.
This motion is a result of interaction between the field and dipole
moment induced in a particle and is called dielectrophoresis
(DEP).
[0010] The dielectrophoretic force on the particle varies with the
frequency of the applied electric field. At the low frequency, the
polarity of the dielectrophoretic force on the particle depends on
the conductivity difference between the particles and electrolyte.
On the other hand, at the high frequency the polarity of the
dielectrophoretic force on the particle depends on the permettivity
difference between the particles and electrolyte. If the particle
is more conductive than the electrolyte around it, the dipole
aligns with the field and the force acts up the field gradient
towards the region of highest electric field. This effect is called
positive dielectrophoresis (PDEP). If the particle is less
polarisable than the electrolyte, the dipole aligns against the
field and the particle is repelled from regions of high electric
field (Hughes, "AC Electrokinetics: Applications for
Nanotechnology", Nanotechnology, 11, pages 124-132, (2000)).This
effect is called negative dielectrophoresis (NDEP).
[0011] Recently, both EP and DEP have captured much interest
because they are effective ways to trap, move, deform and separate
particles ranging from colloidals to DNA strands and biological
cells (Huang, Y; Ewalt, K. L; et al; "Electric Manipulation of
Bioparticles and Macromolecules on Microfabricated electrodes",
Anal Chem, 73, pp. 1549-1559, (2001)). In most cases, embedded
electrodes were carefully designed and fabricated by semiconductor
processing techniques on substrates, such as silicon, glass or
plastics.
[0012] The field-induced assembly method is a unique application of
Electrophoresis/Dielectrophoresis technology. The precise assembly
of two- and three-dimensional colloidal on Conductive ITO electrode
surfaces may be induced by an AC or DC electrical field that is
normal to the electrode surfaces (U.S. Pat. Nos. 5,855,753, and
6,033,547). This technology was extended on silicon electrode, on
which the formation, placement, and rearrangement of planar
colloidal arrays can be effected by an external illumination
pattern due to the photo-assisted impedance modulation. According
to Seul et al, it is necessary to apply an AC electrical field to
penetrate the thin oxide existing on the silicon surface (U.S. Pat.
Nos. 6,251,691, 6,387,707, 6,468,811, 6,514,771, 6,706,163, and
6,797,524). A optoelectronic tweezers has also been demonstrated by
Chiou et al.(Chiou, P. Y; Chang, Z; et al; Proc. IEEE/LEOS
International Conference Optical MEMS, pp. 8-9, (2003)) The
impedance of an amorphous silicon layer, covered by a silicon
nitride layer, is modulated by a laser beam. The particles inside
the electrolyte are polarized by a non-uniform AC field and pushed
away from the illuminated region by the negative dielectrophoresis
force. Those prior arts show superperformances on particle
manipulation, but still have their limitations, such as: 1)
inability to characterize particle electrically; 2) lack of
advantages related to DC electric field; and 3) that the depletion
layer at the semiconductor surface and the polarities switching
with the AC signal make it very hard to precisely control the
electric field applied on the particles.
[0013] Ozkan et al have developed an optical addressing scheme to
localize polymer beads on an unpatterned silicon surface based on a
DC electric field (Ozkan et al, "Heterogeneous Integration through
Electrokinetic Migration", IEEE Engineering in Medicine and
Biology, November/December, pp 144 (2001), or Ozkan et al, "Optical
Addressing of Polymer Beads in Microdevices", Sensor and Materials,
Vol 14, No 4, pp 189-197, (2002)). This approach utilizes an
optical microbeam that is directed on the substrate to create an
active `virtual` electrode (U.S. Pat. No. 6,605,453).The localized
charge is defined by the characteristics of the silicon-electrolyte
interface in the electrochemical system and serves to attract
oppositely charged objects within the solution. Without a layer of
oxide inserted between the silicon and electrolyte, DC voltage was
able to be used to manipulate the particles. This technique also
has its limitations, such as: 1) undesired effects of the dark
current; 2) that high-voltage biasing during the patterning process
must be avoided due to the electrolysis reaction; 3) lack of
advantages from the frequency response of particles on AC field;
and 4) inability to characterize particle electrically.
[0014] In summary, none of the previous efforts in this field
disclose all of the benefits of the present invention, nor does the
prior art teach or suggest all of the elements of the present
invention.
[0015] III. Impedance Spectroscopy
[0016] Electrical impedance spectroscopy (EIS) is widely used in
experimental studies to characterize living cell. For example, EIS
can reflect the size, shape, and density of cells in tissue as well
as the conductivity of intra and extra cellular milieu. This allows
the identification of difference between tissues or between
physiopatological states of the same tissue. The typical way to
perform EIS on samples of tissue is the frequency sweep, with
frequency range from several Hz to several MHz.
[0017] Single cell analysis using DEP and micro electrical
impedance spectroscopy (u-EIS) was demonstrated on bovine
chromaffin cells and red blood cells (Swomitra K, et al, "A Micro
System Dielectrophoresis and Electrical Impedance Spectroscopy for
Cell Manipulation and Analysis", TRANSDUCERS'03, pp 1055-1058,
(2003)). A micro scale electrophysiological analysis system was
fabricated by micromaching technologies and cells were injected
into a microreservoir. Either a vacuum or DEP was utilize to move
cells in the channel and position them between platinum electrodes
for impedance analysis.
[0018] IV. MIS and EIS Tunnel Junction
[0019] The metal-insulator-semiconductor (MIS) structure has been
proved to be extremely useful in semiconductor devices. When an
ideal MIS structure is biased with positive or negative voltages,
four cases may exist at the semiconductor. They are accumulation,
depletion, inversion, and deep depletion cases. (S. M. Sze, "The
Physics of Semiconductor Devices", 2.sup.nd edition, Chapt. 7, pp
362-370, Wiley Interscience (1981))
[0020] Let us use n-type semiconductor as an example. When a
positive voltage is applied to the metal plate, the energy bands
near the semiconductor surface are bent downward. According to
semiconductor theory, the downward bending of the energy bands at
the semiconductor surface gives rise to an enhanced concentration,
an accumulation of electrons near the insulator-semiconductor
interface. This is called the accumulation case.
[0021] When a small negative voltage is applied to the metal
electrode of an ideal MIS structure, the energy bands bend upward.
The majority carriers, electrons here, are pushed away from the
surface by the electric field and depleted at the surface. This is
called the depletion case. The surface region (layer), in which the
majority carriers (electrons) are depleted, is called depletion
region (layer). In the depletion case, the depletion layer will
shield a significant amount of applied electric field.
[0022] According to the semiconductor theory, the hole
concentration at the semiconductor surface is in proportion to the
degree of the upward band bending. When a larger negative bias is
applied, the bands bend upward even more and the hole concentration
at the semiconductor surface may be larger than the intrinsic
carrier concentration and the electron concentration at the surface
becomes less than the intrinsic carrier concentration. The number
of holes (minority carriers) at the surface is greater than the
number of electrons (majority carriers); the surface is thus
inverted. This is called the inversion case and there is an
inversion layer at the insulator-semiconductor interface. As the
band are bent further, eventually the hole concentration at the
surface will be equal to or higher than the original electron
concentration in the n-type semiconductor material. Typically, the
width of the inversion layer ranges from 1 nm to 10 nm and is
always much smaller than the surface depletion layer width. The
inversion layer, after it is formed, will shield most of the
applied electric field and work as a perfect electrode similar to a
piece of metal.
[0023] In addition to the bias condition, the minority carrier
concentration at the insulator-semiconductor interface also depends
on the interaction between the supply capability of minority
carriers and the leakage current through the insulator. Under the
condition that the minority carrier concentration at the
insulator-semiconductor interface is dominated by the leakage
process, the minority carriers (holes for n-type semiconductor)
will leak through the insulator and therefore the inversion layer
can not be formed or maintained at the insulator-semiconductor
interface. The semiconductor surface stays in the depletion region
and this is called the deep depletion case.
[0024] When the thickness of the insulator layer is less than 5nm,
quantum tunneling phenomena plays significant role for a MIS
structure. For a MIS tunnel junction formed on lightly doped
semiconductor in the reverse bias region, the degree of inversion
at semiconductor/ultra-thin insulating layer interface depends on
the supply rate of minority carriers to the surface for a MIS
tunnel junction (Green, M. A; Shewchun, I; Solid-State Electron,
17, pp. 349-365, (1974)). Under the condition without minority
carrier injection, an inversion layer cannot be maintained at the
interface due to the fact that minority carriers leak through
ultra-thin insulating layer, due to the tunneling process. A
significant portion of bias will drop in the depletion region in
the semiconductor and the semiconductor is in the deep depletion
region. With the help of external minority carries injection, such
as illumination, an inversion layer can be built at the interface
and will shield electrical field. As we know, the electron
occupation can be characterized by an energy level, called the
Fermi level, which will change along with applied bias. In the
inversion case, bias will primarily drop on that ultra-thin
insulating layer. As a result, the Fermi level in the metal
electrode will move to majority energy band edge and a majority
carrier tunnel current can be triggered. Theoretically, this
process can result in the multiplication of any minority carrier
current injected to the insulator-semiconductor interface by
factors of 100-1000.
[0025] A localized multiplication process was also observed by this
author in a prior study on a nano-size MIS tunnel junction formed
by a STM tip on lightly doped silicon (Lin, Hai-An et al, Appl.
Phys. Lett. Vol 73 pp. 2462-2464, (1998)). In that case, the
minority carriers were injected by illumination too and MIS tunnel
diodes can work as a photo switch. It has been demonstrated that
current multiplication can occur in a suitably biased MIS tunnel
diode.
[0026] The characterization of electrolyte-insulator-semiconductor
(EIS) junction is very similar to that MIS junction, except the
metal in the MIS junction is replaced by an electrolyte in the EIS
junction. One of well known EIS structures is the
ion-sensitive-filed-transistor (ISFET). The ISFET is constructed by
substituting a sensing film for the metal gate on the gate oxide of
a traditional MOSFET and using electrolyte to apply the gate
voltage. When the thickness of the insulator in an EIS junction is
less than 5 nm, an EIS tunnel junction will be formed.
BRIEF SUMMARY OF THE INVENTION
[0027] The present invention comprises a "sandwich" electrochemical
cell with an inversion layer electrode defined by the area of
illumination region and an instrument to characterize the particles
in the electrolyte. The electrode is formed by an inversion layer
maintained by a proper bias and illumination. With this novel tool,
it is possible to perform biochemical analysis in an integrated
semiconductor chip. In addition, the optoelectronic probe can also
be a powerful tool to assemble next generation nano-electronic or
optoelectronic devices. Basically, this device has the ability to
trap, move, deform, merge, separate and characterize particles,
such as cells, molecules, any type of colloids, any of inorganic
and bio-organic substances, beads, as well as pucks and like small
things on a semiconductor chip.
[0028] ] The first advantage for the present invention is that the
size of the electrode is self-defined by the size of light beam. It
will be easy to adjust the electrode size to match a single cell by
simply adjusting beam size. Therefore we are able to characterize a
single cell in a convenient way. On the other hand, a large beam
size can also be used to assemble a particle array. This is very
different from the Optical Tweezer where the trapping zone is too
small (on the order of the light wavelength) to manipulate the
particle array.
[0029] The second advantage for the present invention is that an
inversion region is used as a working electrode. The inversion
layer has behavior similar to a metal electrode. The most of
applied electric field can be shielded by the inversion layer and
no significant field will penetrate into the semiconductor bulk. In
addition, a majority carrier tunneling process will also reduce
voltage drop on the oxide layer. Therefore a significant portion of
bias will be dropped into the electrolyte. Furthermore, the
metal-like electrode formed by the inversion layer is also perfect
for the impedance characterization. This is very different from the
conventional illumination-assisted field induced assembly
technologies (or a conventional optoelectronic tweezers), where the
semiconductor bulk or depletion region plays as the electrode. The
involvement of the depletion region at the semiconductor surface
will cause an extreme complexity on the voltage distribution among
the different part of the "sandwich" electrochemical cell and make
it very difficulty to analyze the impedance spectroscopy.
[0030] The third advantage for the present invention comes from a
thin oxide layer formed on the silicon surface. This oxide layer
will reduce the dark current significantly and a well defined
electrode can be successfully formed.
[0031] The fourth advantage for the present invention comes from
combination of time constant electric field and time-varying
electric field. The combined use of time constant electric field
and time-varying electric field can complement the limitations of
EP and DEP, and potentially provide an integrated method for
manipulation of bioparticles and macromolecules on microfabricated
chips.
[0032] The fifth advantage for the present invention also comes
from combination of time constant electric field and time-varying
electric field. The combined use of time constant electric field
and time-varying electric field avoids using high DC voltage to
generate high electric field in order to manipulate small
particles. Therefore a strong electrolysis reaction under high DC
current is not going to happen.
[0033] Finally, in the conventional illumination-assisted field
induced assembly technologies (or a conventional optoelectronic
tweezers), the illumination modulates the photoconductivity of
semiconductor. The illumination in the present invention, however,
is only utilized to compensate the leaking minority carrier
current. Therefore, in present invention, one has extreme
flexibility to choose the optical sources and can avoid the
biological object integrity problem caused by focus beam with very
high intensities.
BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWING
[0034] FIG. 1 is a sectional view of an embodiment of the
optoelectronic probe.
[0035] FIG. 2A is the wave function of the time constant
voltage.
[0036] FIG. 2B is the wave function of the DC time-varying
voltage.
[0037] FIG. 2C is the wave function of the combination of the time
constant voltage and DC time-varying voltage.
[0038] FIG. 3 schematically illustrates an embodiment of electric
input, which has the wave function as shown in FIG. 2C.
[0039] FIG. 4A is the energy band diagram for the deep depletion
situation for the EIS Junction.
[0040] FIG. 4B is the energy band diagram for the inversion
situation for the EIS junction.
[0041] FIG. 5 schematically illustrates an embodiment of an
impedance analyzer for characterizing the trapped particles.
DETAILED DESCRIPTION OF THE INVENTION
[0042] The following description is of the best mode presently
contemplated for the carrying out of the invention. This
description is made for the purpose of illustrating the general
principles of the invention, and is not to be taken in a limiting
sense. The scope of the invention is best determined by reference
to the appended claims.
I. Optoelectronic Probe
[0043] FIG. 1 is a sectional view of an embodiment of the present
invention. An optical microscope, represented by reference number
100, can be used to observe and record the particle manipulation
process. The top electrode of the "sandwich" electrochemical cell
is formed by a glass slide 110 coated by an optically transparent
conducting thin film, such as indium tin oxide (ITO) 120. This kind
of transparent conducting electrode is commercially available. A
spacer (typically thick .about.50 .mu.m), represented by reference
number 130, is formed by polymer film with a hole in the center.
Reference numeral 140 denotes a layer of thin oxide (typically
about 15-30 Angstroms thick) and reference numeral 150 denotes a
piece of n-type silicon, with doping density range from
10.sup.16-10.sup.17 cm.sup.-3. The properly doped n-type silicon,
covered with a thin oxide layer, is used to form the bottom
electrode, where an Electrolyte/Insulator/Semiconductor (EIS)
structure will perform a Metal-Insulator-Semiconductor-like, or
MIS-like tunnel diode. Reference numeral 160 denotes the
electrolyte. The bottom electrode will also work as a stage to
support particles, which are represented by reference number 170.
Contact to electrodes is achieved through leads in the form of Au
wires by means of silver epoxy. Reference numeral 180 denotes a
light beam from the illumination source and reference numeral 190
denotes an inversion region, which works as a working electrode to
control the movement of particles in the electrolyte. A laser beam,
such as a 633 nm He--Ne laser, or IR laser diode, range from 650 to
680 nm, can be used here as a illumination source to inject
minority carriers. The laser beam is steered by a pair of
orthogonally scanning galvanometer mirrors as described in a paper
by P. Y. Chiou et al. range (Chiou, P. Y; Chang, Z; et al; Proc.
IEEE/LEOS International Conference Optical MEMS, pp. 8-9, (2003)).
The beam is sent through a microscope and the objective lens is
used to both focus the beam and view the trapped object. For
assembling a particle array, a regular microscope illuminator can
also be used here, with apertures or masks to provide the required
shape or size of beam (Seul, M et al, U.S. Pat. No. 6,251,691
(1997)). In addition, a CCD camera is used to monitor the trapped
objects.
[0044] The silicon surfaces are carefully cleaned in adherence with
semiconductor industry standard RCA and Piranha cleaning protocols.
The original "native" oxide can be removed by HF solution. The thin
oxide, represented by reference number 140 as shown in FIG. 1, can
be thermally grown under standard conditions in a furnace at 950
degrees C. In addition, the thin oxide layers can also be regrown
after removal of the original "native" oxide in HF solution, under
UV illumination from a deuterium source in the presence of oxygen.
The back side oxide is stripped away, using conventional oxide
stripping process. In order to have better performance, silicon 150
in the FIG. 1 can be replaced by a piece of epitaxial silicon. The
n.sup.- (.rho..about.15-20 .OMEGA.-cm) epi layer about 6-8 .mu.m
will make sure a depletion layer at the silicon surface and 200-250
.mu.m thick n.sup.+ (.rho..about.0.01 .OMEGA.-cm) substrate is used
to reduce the serial resistance. All kinds of silicon wafers are
commercially available throughout the semiconductor industry.
II. Electric Input
[0045] A positive time constant bias is denoted by reference number
200 in FIG. 2A. A time-varying voltage with chosen frequency is
denoted by reference number 210 in FIG. 2B. In order to make sure
semiconductor surface stay either in the depletion region or strong
inversion region, we are using a DC time-varying voltage signal
here, as shown in FIG. 2B. The DC time-varying voltage 210 will
superpose to the positive time constant bias 200. The final applied
input is denoted by reference number 220, as shown in FIG. 2C.
[0046] FIG. 3 schematically illustrates an embodiment of electric
input 220 shown in FIG. 2C. The time constant voltages are
generated by a commercially available voltage source 300, with
applied bias adjustable. The time-varying voltages are produced by
a function generator 310, with frequencies varying from DC to
several MHz. A commercially available full wave rectifier 320 is
used to rectify the AC input signal from the function generator
into a DC time varying voltage signal. The positive time constant
voltages, superposed by the DC time-varying voltages, are applied
to the silicon piece through terminal 330. In this way, the n-type
silicon electrode will always stay in the positive bias region. Due
to current leakage via tunneling between the red-ox level
E.sub.redox in the electrolyte and the valence band of the silicon,
the thin oxide/silicon interface will be set into deep depletion
region by the applied bias and the depletion layer at the
semiconductor surface will shield most of the external electrical
field. It is just like a switch being turned off and no significant
electrical field is able to penetrate into the electrolyte.
III. Inversion Layer Electrode
[0047] The energy band diagram for the deep depletion situation is
shown in FIG. 4A. The red-ox level E.sub.redox is represented by
reference number 400. The potential barrier formed by ultra-thin
oxide is denoted by reference number 410. Reference number 420
represents the conduction band edge, reference number 430
represents the valence band edge and reference number 440
represents the silicon Fermi level. The reference number 450
represents the depletion region at the silicon surface. In order to
simply the discussion, here we ignore the space charge region in
the electrolyte.
[0048] If a beam of light injects into the surface of deep depleted
EIS tunnel diode, extra holes, which are the minority carrier
herein, will be supplied to the silicon surface to compensate the
leaked holes. The semiconductor surface in the illumination area
will be turned into inversion. The holes, denoted by the reference
number 460 in FIG. 4B, accumulate at the surface and work as an
electrode to shield the electrical field, as denoted by the
reference number 190 in FIG. 1. Further more, the red-ox level 400
in the electrolyte will be moved up above the semiconductor
conduction band 420, as shown in FIG. 4B and, a majority carrier
current can be carried by direct tunneling process between the
red-ox level and conduction band. Therefore, a significant bias
will be dropped in the electrolyte in the illuminated area. Outside
the illuminated area, the semiconductor still stay in the deep
depletion and the electric field is shielded by the depletion
layer. Since the inversion layer electrode 190 has area much
smaller than that of ITO film 120, as shown in FIG. 1, an electric
field concentration region has been well defined by the illuminated
area and a non-uniform electric field has been built into the
electrolyte between the top electrode 120 and the bottom electrode
190 formed by the inversion layer.
IV. Basic Operations
[0049] The non-uniform electric field can be used to trap charged
particles by EP process or polarized (charged or neutral) particles
by DEP process. An array of particles can be assembled in a
designed area, and the interparticle spacing and internal state of
order within the array may be controlled by adjusting the applied
field prior to anchoring the array to the substrate.
[0050] An array of particles starts with a capture process. As long
as the inversion layer electrode is formed, particles will be
captured over it. An array starts to grow and will continue to grow
until it approaches the outer limits of the electrode. The internal
state of order of captured aggregate of particles is determined by
the strength of applied voltage, high values favoring increasingly
denser pacing of particles and the eventual formation of ordered
arrays displaying a hexagonally crystalline configuration in the
form of a bubble raft. The size and shape of the array are
determined by the light beam. Arrays are maintained by the applied
electric field in a liquid environment. Removal of the applied
voltage results in the disassembly of the array.
[0051] The process also leaves the array in a state that may be
readily subjected to further chemical modification such as
cross-linking, or made permanent by chemical anchoring to the
substrate. This is best accomplished by involving anchoring
chemistries analogous to those relying on heterbifunctional
cross-linking agents invoked to anchor proteins via amide bond
formation. Molecular recognition, for example between biotinylated
particles and surface-anchored streptavidin, provides another class
of coupling chemistries for permanent anchoring.
[0052] If the light beam moves from one position to the next
position, the inversion layer electrode will move along with the
beam and so will the trapped particle array, as long as the light
beam has a moving speed slower than that of particles, dragged by
the lateral component of the non-uniform electric field. The
control of lateral particle transport by changing or moving
patterns of illumination has the advantage that it may be applied
whenever and wherever required.
V. Time Constant Fields
[0053] Under the condition that only a time constant voltage is
used, the polarity of the dielectrophoretic force on the particle
depends on the conductivity difference between the particles and
electrolyte. If the particle is more conductive than the
electrolyte around it, the dipole aligns with the field and the
force acts up the field gradient towards the region of highest
electric field. Examples of trapping polarized (charged or neutral)
particles by DEP process can be demonstrated by colloidal beads,
such as silica, and polystyrene, with a diameter in the range from
several hundred Angstroms to 2 .mu.m. With silica, the electrolyte
is triply distilled water (pH=5.8 with a conductivity of <1
.mu.S cm.sup.-1); for polystyrene the suspending electrolyte
contained a mixture of ionic and nonionic surfactants added to
provide the colloidal stability. At proper electric field range
(<100 V cm.sup.-1 typically), the particle trapping process is
reversible. When the field is removed, the particle array is
stirred by Brownian motion. On the other hand, a strong electric
field compresses the particles and they coagulate or adhere to the
electrode surface. The array will be assembled and permanently
frozen on the electrode surface. With properly adjusting the size
of illumination (focused laser beam), we can have a reasonable size
of electrode and therefore all the operations mentioned above can
be repeated on a single particle, or a single living cell.
VI. Combination of Time Constant/Time-Varying Fields
[0054] When a time-varying voltage is superposed to the applied
time constant voltage, the combination of the time-varying and time
constant electric field will bring us extreme flexibility to
manipulate particles. This is because that the dielectrophoretic
force on the particle will vary with the frequency of the applied
electric field. If the particle is less polarisable than the
electrolyte, the dipole aligns against the field and the particle
is repelled from regions of high electric field. This effect is
called negative dielectrophoresis (NDEP). Particles having
different dielectric properties will experience different DEP
forces that may be exploited for particle selective manipulation.
Therefore, the frequency of the time-varying voltage can be chosen
to selectively trap or deform particles if they are deformable
particles, such as living cells.
[0055] This example will show how we can separate spores to
bacteria. Sample preparation process is similar to one described in
a paper by Y. Huang et al. (Huang, Y; Ewalt, K. L; et al; "Electric
Manipulation of Bioparticles and Macromolecules on Microfabricated
electrodes", Anal Chem, 73, pp. 1549-1559, (2001)). B. globigii
spores (the Biological Defense Research Directorate, Bethesda, Md.)
were stocked in phosphate-buffered saline (PBS, pH=7.2; Life
Technologies, Grand Island, N.Y.) at a concentration of
1.4.times.10.sup.9 spores/mL. Heat-killed E. coli 0157:H7 bacteria
(KPL, Gaithersburg, Md.) were stocked in distilled water at a
concentration of 7.times.10.sup.9 bacteria/mL. Heat-killed E. coli
bacteria and B. globigii spores were washed 3 times in 280 mM
mannitol (Sigma) nd resuspended in the mannitol solution having a
conductivity of 20 .mu.S/cm. The final mixture concentration was
2.times.10.sup.8/mL and 3.times.10.sup.8 mL for bacteria and
spores, respectively. The time-varying voltage here used has
frequency about 50 KHz. The bacteria were repelled from the
inversion layer electrode by NDEP force and the spores were trapped
over the electrode by DEP force. If the light beam moves from one
position to the next position, the trapped spores will moves along
with the beam. Therefore, the spores will be separated further from
the bacteria.
[0056] The negative DEP force also gives us another way to
transport particles in the electrolyte on the semiconductor
surface. A 100 KHz time-varying voltage is applied in this case to
drive 20 .mu.m polystyrene particles (from Polysciences). The
buffer solution consists of deionized water and KCL, mixed to have
a conductivity of 10 mS/m. The particle is pushed by the optical
beam, until at sufficiently high scan rate.
VII. Impedance Spectroscopy
[0057] Conventional impedance-based sensing of particles is a
well-accepted method for the counting, sizing and characterizing of
particles and cells and finds wide application in clinical and
veterinary laboratories for the analysis of blood, cell
suspensions, and other samples. In general, the particle response
is frequency dependent and the frequency dependency can be used to
characterize or identify the particle. Applications of the present
invention are vast and include, but are not limited to applications
such as cell and particle counting, cell and particle subpopulation
analysis, cell viability analysis, cell and particle concentration
analysis, cell differential analysis, medical applications,
veterinary applications, bioengineering, food analysis, soil
analysis, in-line particle detection in fluidic circuits and
systems, detection of bacterial spores and other biological agents
of potential use in warfare and terrorism, discrimination of
potentially harmful biological agents from non harmful biological
cells such as pollen and from inert particulate materials such as
dust, smoke, and non-viable cells, detection of responses of cells
such as human blood cell subpopulations to biological and chemical
agents, and detection and discrimination of bacterial cells and
spores (including anthrax) for medical, agricultural,
environmental, and bio-warfare and bio-terrorism detection
applications.
[0058] FIG.5 schematically illustrates an embodiment of an
impedance analyzer for characterizing the trapped particles in the
field concentration region on the bottom silicon electrode 150 of
FIG. 1. As shown in the FIG. 5, an impedance analyzer consists of a
digital oscilloscope 500 to sense the current signal and a personal
computer 510 to analyze the current signal and generate the
impedance spectroscopy. The data bus 520 is responsible for the
communication between the digital oscilloscope 500 and personal
computer 510. The input of the digital oscilloscope 500 is
connected to top ITO electrode 120 of FIG. 1 to collect the current
signal. The terminal 530, which in one side is contact to the
bottom silicon electrode 150 of FIG. 1, is connected to terminal
330 in FIG. 3. The out put of the rectifier 490 in FIG. 3 gives us
the desired voltage signal as shown in FIG. 2C. An array of
particles can be characterized by the impedance analyzed after it
is assembled. A classical method to perform impedance measurements
on particles is the frequency sweep, with the frequency range of
the time variable electrical field 310 in FIG. 3 from several Hz to
several MHz. The responses of particles on applied electric fields
are sensed by the digital oscilloscope 500 and analyzed by the
personal computer 510. Both the magnitude and phase measurements of
the impedance have interesting implications, in regards to particle
characterization and identification.
[0059] In the case that a single cell is trapped by the
optoelectronic probe, we are able to characterize the single cell
by the impedance analyzer. Since the inversion layer electrode is
defined by the illumination, it will be easy to make it match the
cell size by simply adjusting beam size. In addition, the inversion
layer will shield most of the applied electric field and work as a
perfect electrode similar to a metal piece. This is very different
from the conventional illumination-assisted field induced assembly
technologies (or a conventional optoelectronic tweezers), where the
semiconductor bulk, or depletion region plays as the electrode. The
involvement of the depletion region at the semiconductor surface
will cause an extreme complexity on the voltage distribution among
the different part of the "sandwich" electrochemical cell and make
it very difficulty to analysis the impedance spectroscopy. All
those features in the present invention will give us an edge to
sense the single cell.
VIII. Conclusion
[0060] All of the methods disclosed and claimed herein can be made
and executed without undue experimentation in light of the present
disclosure. While the techniques of this invention have been
described in terms of specific embodiments, it will be apparent to
those of skill in the art that variations may be applied to the
methods and in the steps or in the sequence of steps of the method
described herein without departing from the concept, spirit and
scope of the invention. All such similar substitutes and
modifications apparent to those skilled in the art are deemed to be
within the spirit, scope and concept of the invention as defined by
the appended claims.
* * * * *