U.S. patent application number 10/979054 was filed with the patent office on 2005-08-11 for electrochemical methods and devices for use in the determination of hematocrit corrected analyte concentrations.
This patent application is currently assigned to LifeScan, Inc. Invention is credited to Kermani, Mahyar Z., O'hara, Timothy.
Application Number | 20050176153 10/979054 |
Document ID | / |
Family ID | 23976305 |
Filed Date | 2005-08-11 |
United States Patent
Application |
20050176153 |
Kind Code |
A1 |
O'hara, Timothy ; et
al. |
August 11, 2005 |
Electrochemical methods and devices for use in the determination of
hematocrit corrected analyte concentrations
Abstract
Methods and devices for determining the concentration of an
analyte in a physiological sample are provided. In the subject
methods, the physiological sample is introduced into an
electrochemical cell having a working and reference electrode. A
first electric potential is applied to the cell and the resultant
cell current over a period of time is measured to determine a first
time-current transient. A second electric potential of opposite
polarity is then applied and a second a time-current transient is
determined. The preliminary concentration of the analyte is then
calculated from the first and/or second time-current transient.
This preliminary analyte concentration less a background value is
then multiplied by a hematocrit correction factor to obtain the
analyte concentration in the sample, where the hematocrit
correction factor is a function of the preliminary analyte
concentration and the variable .gamma. of the electrochemical cell.
The subject methods and devices are suited for use in the
determination of a wide variety of analytes in a wide variety of
samples, and are particularly suited for the determination of
analytes in whole blood or derivatives thereof, where an analyte of
particular interest is glucose.
Inventors: |
O'hara, Timothy; (Danville,
CA) ; Kermani, Mahyar Z.; (Pleasanton, CA) |
Correspondence
Address: |
BOZICEVIC, FIELD & FRANCIS LLP
1900 UNIVERSITY AVENUE
SUITE 200
EAST PALO ALTO
CA
94303
US
|
Assignee: |
LifeScan, Inc
Milpitas
CA
|
Family ID: |
23976305 |
Appl. No.: |
10/979054 |
Filed: |
November 1, 2004 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10979054 |
Nov 1, 2004 |
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10144095 |
May 10, 2002 |
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6890421 |
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10144095 |
May 10, 2002 |
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09497304 |
Feb 2, 2000 |
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6475372 |
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Current U.S.
Class: |
436/70 ;
204/450 |
Current CPC
Class: |
G01N 27/3274
20130101 |
Class at
Publication: |
436/070 ;
204/450 |
International
Class: |
G01N 027/447 |
Claims
1-20. (canceled)
21. An electrochemical cell for determining the concentration of an
analyte in a physiological sample, the electrochemical cell
comprising: a working electrode; a reference electrode; a spacer
layer between the working and reference electrodes; and a reaction
zone defined by the spacer layer and the working and reference
electrodes, said reaction zone comprising an enzyme for producing
an electrochemical signal in the presence of the analyte, wherein
the reaction zone has a volume of from about 0.1 .mu.l to about 10
.mu.l; wherein the electrochemical cell provides a measurement that
correlates with the concentration of the analyte in the
physiological sample within a period from about 3 to about 20
seconds.
22. The electrochemical cell of claim 21, wherein the reaction zone
of from about 0.9 .mu.l to about 1.6 .mu.l;
23. The electrochemical cell of claim 21, wherein the period is
from about 4 to about 10 seconds.
24. The electrochemical cell of claim 21, wherein the analyte is
glucose and the enzyme is glucose oxidase.
25. The electrochemical cell of claim 1, wherein said reaction zone
further comprises a mediator.
26. The electrochemical cell of claim 25, wherein said mediator is
ferricyanide.
27. A system for determining the concentration of an analyte in a
physiological sample, the system comprising: a test strip
comprising the electrochemical cell of claim 21; and a meter for
receiving the test strip, the meter comprising means for applying a
first constant electric potential between the working and the
reference electrodes over a first period of time to obtain a first
time-current transient and for applying a second constant electric
potential between the working and the reference electrodes over a
second period of time to obtain a second time-current transient,
and comprising means for deriving an analyte concentration from
said first and second time-current transients.
28. A method for determining the concentration of an analyte in a
physiological sample, the method comprising: providing the
electrochemical cell of claim 1; introducing the sample into the
reaction zone of the electrochemical cell; applying a constant
electric potential between the working and the reference
electrodes; observing a change in current between the electrodes
over first period of time from about 3 to about 20 seconds to
obtain a first time-current transient; applying a second constant
electric potential between the working and the reference
electrodes; observing a change in current between the electrodes
over a second period of time from about 1 to about 10 seconds to
obtain a second time-current transient; and deriving an analyte
concentration from said first and second time-current
transients.
29. The method of claim 28 wherein the total amount of time
required to obtain the first and second time-current transients is
less than about 30 seconds.
30. The method of claim 29 wherein the total amount of time
required to obtain the first and second time-current transients is
less than about 20 seconds.
31. The method of claim 30 wherein the total amount of time
required to obtain the first and second time-current transients is
less than about 14 seconds.
32. The method of claim 28 wherein the magnitude of the first
applied electric potential is opposite that of the magnitude of the
second applied electric potential.
33. The method of claim 28 wherein the magnitude of the first
applied electric potential is from about 0 to about -0.6 V and the
second constant electric potential typically ranges from about 0 to
about +0.6 V.
34. The method of claim 33 wherein the magnitude of the first
applied electric potential is from about -0.2 to -0.4 V and the
second constant electric potential typically ranges from about +0.2
to +0.4 V.
Description
FIELD OF THE INVENTION
[0001] The field of this invention is analyte determination,
particularly electrochemical analyte determination and more
particularly the electrochemical determination of blood
analytes.
BACKGROUND
[0002] Analyte detection in physiological fluids, e.g. blood or
blood derived products, is of ever increasing importance to today's
society. Analyte detection assays find use in a variety of
applications, including clinical laboratory testing, home testing,
etc., where the results of such testing play a prominent role in
diagnosis and management in a variety of disease conditions.
Analytes of interest include glucose for diabetes management,
cholesterol, and the like. In response to this growing importance
of analyte detection, a variety of analyte detection protocols and
devices for both clinical and home use have been developed.
[0003] One type of method that is employed for analyte detection is
an electrochemical method. In such methods, an aqueous liquid
sample is placed into a reaction zone in an electrochemical cell
comprising two electrodes, i.e. a reference and working electrode,
where the electrodes have an impedance which renders them suitable
for amperometric measurement. The component to be analyzed is
allowed to react directly with an electrode, or directly or
indirectly with a redox reagent to form an oxidisable (or
reducible) substance in an amount corresponding to the
concentration of the component to be analyzed. i.e. analyte. The
quantity of the oxidisable (or reducible) substance present is then
estimated electrochemically and related to the amount of analyte
present in the initial sample.
[0004] Where the physiological sample being assayed is whole blood
or a derivative thereof, the hematocrit of the sample can be a
source of analytical error in the ultimate analyte concentration
measurement. For example, in electrochemical measurement protocols
where the analyte concentration is derived from observed
time-current transients, hematocrit can slow the equilibration
chemistry in the electrochemical cell and/or slow the enzyme
kinetics by increasing the sample viscosity in the cell, thereby
attenuating the time current response and causing analytical
error.
[0005] As such, there is great interest in the development of
methods of at least minimizing the hematocrit originated analytical
error. In certain protocols, blood filtering membranes are employed
to remove red blood cells and thereby minimize the hematocrit
effect. These particular protocols are unsatisfactory in that
increased sample volumes and testing times are required. Other
protocols focus on the determination of the capillary fill time.
However, these protocols add complexity to both the strips and
devices that are used to read them. In yet other embodiments,
hematocrit is separately determined using two additional
electrodes, which also results in more complex and expensive
strips/devices.
[0006] As such, there is continued interest in the identification
of new methods for electrochemically measuring the concentration of
an analyte in a physiological sample, where the method minimizes
the analytical error which originates with the hematocrit of the
sample.
[0007] Relevant Literature
[0008] Patent documents of interest include: U.S. Pat. No.
5,942,102 and WO 97/18465.
SUMMARY OF THE INVENTION
[0009] Methods and devices for determining the concentration of an
analyte in a physiological sample are provided. In the subject
methods, the physiological sample is introduced into an
electrochemical cell having a working and reference electrode. A
first electric potential is applied to the cell and the resultant
cell current over a first period of time is measured to determine a
first time-current transient. A second electric potential of
opposite polarity is then applied to the cell and a second
time-current transient is determined. The preliminary concentration
of the analyte (C.sub.0) is then calculated from the first and/or
second time-current transients. This preliminary analyte
concentration, less a background value, is then multiplied by a
hematocrit correction factor to obtain the analyte concentration in
the sample, where the hematocrit correction factor is a function of
the preliminary analyte concentration and the ratio of 2 current
values (.gamma.) within the time-current transient of the
electrochemical cell. The subject methods and devices are suited
for use in the determination of a wide variety of analytes in a
wide variety of samples, and are particularly suited for the
determination of analytes in whole blood or derivatives thereof,
where an analyte of particular interest is glucose.
BRIEF DESCRIPTION OF THE FIGURES
[0010] FIG. 1 provides a three-dimensional graph of C.sub.0,
.gamma. and .alpha.(C.sub.0, .gamma.) derived from experimental
data using a wide range of glucose and hematocrit values.
DESCRIPTION OF THE SPECIFIC EMBODIMENTS
[0011] Methods and devices for determining the concentration of an
analyte in a physiological sample are provided. In the subject
methods, the physiological sample is introduced into an
electrochemical cell having a working and reference electrode. A
first electric potential is applied to the cell and the resultant
cell current over a first period of time is measured to determine a
first time-current transient. A second electric potential of
opposite polarity is then applied to the cell and a second a
time-current transient is determined. The preliminary concentration
of the analyte is then calculated from the first and/or second
time-current transient. This preliminary analyte concentration,
less a background value, is then multiplied by a hematocrit
correction factor to obtain the analyte concentration in the
sample, where the hematocrit correction factor is a function of the
preliminary analyte concentration and the variable .gamma. of the
electrochemical cell. The subject methods and devices are suited
for use in the determination of a wide variety of analytes in a
wide variety of samples, and are particularly suited for use in the
determination of analytes in whole blood or derivatives thereof,
where an analyte of particular interest is glucose. In further
describing the subject invention, the subject methods will be
described first followed by a review of a representative device for
use in practicing the subject methods.
[0012] Before the subject invention is described further, it is to
be understood that the invention is not limited to the particular
embodiments of the invention described below, as variations of the
particular embodiments may be made and still fall within the scope
of the appended claims. It is also to be understood that the
terminology employed is for the purpose of describing particular
embodiments, and is not intended to be limiting. Instead, the scope
of the present invention will be established by the appended
claims.
[0013] In this specification and the appended claims, singular
references include the plural, unless the context clearly dictates
otherwise. Unless defined otherwise, all technical and scientific
terms used herein have the same meaning as commonly understood to
one of ordinary skill in the art to which this invention
belongs.
[0014] Methods
[0015] As summarized above, the subject invention provides a method
for determining a hematocrit corrected analyte concentration value
in a physiological sample. By hematocrit corrected analyte
concentration is meant that the analyte concentration value
determined using the subject methods has been modulated or changed
to remove substantially all contribution of hematocrit to the
value. In other words, the concentration value that is determined
using the subject methods has been modified so that any
contribution to the value from the hematocrit of the sample that
would be present in the value but for the practicing of the subject
methods is removed. As such, the hematocrit signal is deconvoluted
from the analyte signal in the subject methods, and only the
analyte signal is employed in arriving at the final hematocrit
corrected analyte concentration.
[0016] The first step in the subject methods is to introduce a
quantity of the physiological sample of interest into an
electrochemical cell that includes spaced apart working and
reference electrodes and a redox reagent system. The physiological
sample may vary, but in many embodiments is generally whole blood
or a derivative or fraction thereof, where whole blood is of
particular interest in many embodiments. The amount of
physiological sample, e.g. blood, that is introduced into the
reaction area of the test strip varies, but generally ranges from
about 0.1 to 10 .mu.L, usually from about 0.9 to 1.6 .mu.L. The
sample is introduced into the reaction area using any convenient
protocol, where the sample may be injected into the reaction area,
allowed to wick into the reaction area, and the like, as may be
convenient.
[0017] While the subject methods may be used, in principle, with
any type of electrochemical cell having spaced apart working and
reference electrodes and a redox reagent system, in many
embodiments the subject methods employ an electrochemical test
strip. The electrochemical test strips employed in these
embodiments of the subject invention are made up of two opposing
metal electrodes separated by a thin spacer layer, where these
components define a reaction area or zone in which is located a
redox reagent system.
[0018] In certain embodiments of these electrochemical test strips,
the working and reference electrodes are generally configured in
the form of elongated rectangular strips. Typically, the length of
the electrodes ranges from about 1.9 to 4.5 cm, usually from about
2.0 to 2.8 cm. The width of the electrodes ranges from about 0.38
to 0.76 cm, usually from about 0.51 to 0.67 cm. The reference
electrodes typically have a thickness ranging from about 10 to 100
nm and usually from about 10 to 20 nm. In certain embodiments, the
length of one of the electrodes is shorter than the length of the
other electrode, typically about 0.32 cm. The shorter electrode may
be the working or reference electrode.
[0019] The working and reference electrodes are further
characterized in that at least the surface of the electrodes that
faces the reaction area in the strip is a metal, where metals of
interest include palladium, gold, platinum, silver, iridium,
carbon, doped tin oxide, stainless steel and the like. In many
embodiments, the metal is gold or palladium. While in principle the
entire electrode may be made of the metal, each of the electrodes
is generally made up of an inert support material on the surface of
which is present a thin layer of the metal component of the
electrode. In these more common embodiments, the thickness of the
inert backing material typically ranges from about 51 to 356 .mu.m,
usually from about 102 to 153 .mu.m while the thickness of the
metal layer typically ranges from about 10 to 100 nm and usually
from about 10 to 40 nm, e.g. a sputtered metal layer. Any
convenient inert backing material may be employed in the subject
electrodes, where typically the material is a rigid material that
is capable of providing structural support to the electrode and, in
turn, the electrochemical test strip as a whole. Suitable materials
that may be employed as the backing substrate include plastics,
e.g. PET, PETG, polyimide, polycarbonate, polystyrene, silicon,
ceramic, glass, and the like.
[0020] A feature of the electrochemical test strips used in these
embodiments of the subject methods is that the working and
reference electrodes as described above face each other and are
separated by only a short distance, such that the distance between
the working and reference electrode in the reaction zone or area of
the electrochemical test strip is extremely small. This minimal
spacing of the working and reference electrodes in the subject test
strips is a result of the presence of a thin spacer layer
positioned or sandwiched between the working and reference
electrodes. The thickness of this spacer layer generally should be
less than or equal to 500 .mu.m, and usually ranges from about 102
to 153 .mu.m. The spacer layer is cut so as to provide a reaction
zone or area with at least an inlet port into the reaction zone,
and generally an outlet port out of the reaction zone as well. The
spacer layer may have a circular reaction area cut with side inlet
and outlet vents or ports, or other configurations, e.g. square,
triangular, rectangular, irregular shaped reaction areas, etc. The
spacer layer may be fabricated from any convenient material, where
representative suitable materials include PET, PETG, polyimide.
polycarbonate, and the like, where the surfaces of the spacer layer
may be treated so as to be adhesive with respect to their
respective electrodes and thereby maintain the structure of the
electrochemical test strip. Of particular interest is the use of a
die-cut double-sided adhesive strip as the spacer layer.
[0021] The electrochemical test strips used in these embodiments of
the subject invention include a reaction zone or area that is
defined by the working electrode, the reference electrode and the
spacer layer, where these elements are described above.
Specifically, the working and reference electrodes define the top
and bottom of the reaction area, while the spacer layer defines the
walls of the reaction area. The volume of the reaction area is at
least about 0.1 .mu.L, usually at least about 1 .mu.L and more
usually at least about 1.5 .mu.L, where the volume may be as large
as 10 .mu.L or larger. As mentioned above, the reaction area
generally includes at least an inlet port, and in many embodiments
also includes an outlet port. The cross-sectional area of the inlet
and outlet ports may vary as long as it is sufficiently large to
provide an effective entrance or exit of fluid from the reaction
area, but generally ranges from about 9.times.10.sup.-4 to
5.times.10.sup.-3 cm.sup.2, usually from about 1.3.times.10.sup.-3
to 2.5.times.10.sup.-3cm.sup.2.
[0022] Present in the reaction area is a redox reagent system,
which reagent system provides for the species that is measured by
the electrode and therefore is used to derive the concentration of
analyte in a physiological sample. The redox reagent system present
in the reaction area typically includes at least an enzyme(s) and a
mediator. In many embodiments, the enzyme member(s) of the redox
reagent system is an enzyme or plurality of enzymes that work in
concert to oxidize the analyte of interest. In other words, the
enzyme component of the redox reagent system is made up of a single
analyte oxidizing enzyme or a collection of two or more enzymes
that work in concert to oxidize the analyte of interest. Enzymes of
interest include oxidases, dehydrogenases, lipases, kinases,
diphorases, quinoproteins, and the like.
[0023] The specific enzyme present in the reaction area depends on
the particular analyte for which the electrochemical test strip is
designed to detect, where representative enzymes include: glucose
oxidase, glucose dehydrogenase, cholesterol esterase, cholesterol
oxidase, lipoprotein lipase, glycerol kinase, glycerol-3-phosphate
oxidase, lactate oxidase, lactate dehydrogenase, pyruvate oxidase,
alcohol oxidase, bilirubin oxidase, uricase, and the like. In many
preferred embodiments where the analyte of interest is glucose, the
enzyme component of the redox reagent system is a glucose oxidizing
enzyme. e.g. a glucose oxidase or glucose dehydrogenase.
[0024] The second component of the redox reagent system is a
mediator component, which is made up of one or more mediator
agents. A variety of different mediator agents are known in the art
and include: ferricyanide, phenazine ethosulphate, phenazine
methosulfate, pheylenediamine, 1-methoxy-phenazine methosulfate,
2,6-dimethyl-1,4-benzoquinone, 2,5-dichloro-1,4-benzoquinone,
ferrocene derivatives, osmium bipyridyl complexes, ruthenium
complexes, and the like. In those embodiments where glucose in the
analyte of interest and glucose oxidase or glucose dehydrogenase
are the enzyme components, mediators of particular interest are
ferricyanide, and the like.
[0025] Other reagents that may be present in the reaction area
include buffering agents, e.g. citraconate, citrate, malic, maleic,
phosphate, "Good" buffers and the like. Yet other agents that may
be present include: divalent cations such as calcium chloride, and
magnesium chloride; pyrroloquinoline quinone; types of surfactants
such as Triton, Macol, Tetronic, Silwet, Zonyl, and Pluronic;
stabilizing agents such as albumin, sucrose, trehalose, mannitol,
and lactose.
[0026] The redox reagent system is generally present in dry form.
The amounts of the various components may vary, where the amount of
enzyme component typically ranges from about 1 to 100 mg/mL,
usually from about 5 to 80 mg/mL; and the amount of mediator
component typically ranges from about 5 to 1000 mM, usually from
about 90 to 900 mM.
[0027] Following sample introduction, first and second time-current
transients are obtained. The first and second time-current
transients are obtained by applying a constant electric potential
to the cell and observing the change in current over a period of
time in the cell. In other words, first and second pulses are
applied to the cell and the resultant time-current transients are
observed. As such, the first time-current transient is obtained by
applying a constant electric potential or first pulse to the cell,
e.g. between the working and the reference electrodes, and
observing the change in current over time between the electrodes,
i.e. change in cell current, to obtain the first time-current
transient. The magnitude of the first applied electric potential
generally ranges from about 0 to -0.6 V, usually from about -0.2 to
-0.4 V. The length of time over which the current between the
electrodes is observed to obtain the first time-current transient
typically ranges from about 3 to 20 seconds, usually from about 4
to 10 seconds.
[0028] The second time current is obtained by applying a second
constant electric potential or second pulse, typically of opposite
polarity from the first constant electric potential, to the
electrodes and observing the change in current between the
electrodes for a second period of time. The magnitude of this
second constant electric potential typically ranges from about 0 to
+0.6 V, usually from about +0.2 to +0.4 V, where in many
embodiments the magnitude of the second electric potential is the
same as the magnitude of the first electric potential. The second
time period typically ranges from about 1 to 10 seconds, usually
from about 2 to 4 seconds. By observing the change in current
between the electrodes over this second period of time, a second
time-current transient for the cell is determined.
[0029] The overall time period required to obtain the requisite
first and second time-current transients, as described above, is
relatively short in certain embodiments. In such embodiments, the
total amount of time required to obtain the first and second
time-current transients is less than about 30 seconds, usually less
than about 20 seconds and more usually less than about 14
seconds.
[0030] The next step in the subject methods is to use the observed
first and second time-current transients, obtained as described
above, to determine: (a) the variable .gamma. of the
electrochemical cell used in the subject methods; and (b) a
preliminary analyte concentration for the analyte of interest in
the sample.
[0031] The variable .gamma. employed in the subject methods is
defined to describe the deviation of the electrochemical cell from
ideality. By way of background, it should be noted that .gamma.
should approach unity under ideal conditions, i.e. reagent
equilibration and glucose reaction are complete before the end of
the first pulse. Any of these conditions not being complete will
cause the ratio to deviate fron non-unity values. The numerator of
.gamma. is defined as the steady-state current observed following
application of the second electric potential to the cell, i.e.
predicted value at t=.infin. of the second time-current transient.
The denominator is defined as the average current over a short time
period near the end of the first period of time, i.e. near the end
of the application of the first electric potential or first pulse.
The short period of time from which the average current is
determined typically ranges from 0.2 to 2 seconds, usually from
about 0.2 to 1.5 seconds and more usually from about 0.2 to 1.25
seconds, where in many embodiments the short period of time is
about 0.3 second. The average current is determined at a time near
the end of the first time period, typically within about 0.1 to 1
second. In certain embodiments, the variable .gamma. is described
by the formula:
.gamma.=i.sub.ss/i.sub.pp
[0032] where:
[0033] i.sub.ss is the steady-state current of the second applied
electric potential; and
[0034] i.sub.pp is the average current over a short period of time
near the end of first time period, i.e. near the end of the time
during which the first electric potential is applied to the cell.
For example, where the first time period is 10 seconds long, the
average current may be the average current from 8.5 to 9.5 seconds
of the 10 second long period, which is a 1.0 second time period 0.5
seconds from the end of the first time period As mentioned above,
the first and second time-current transients are also employed to
derive a preliminary analyte concentration value for the sample
being assayed. In many embodiments, the preliminary analyte
concentration is determined by using the following equations:
i(t)=i.sub.ss{1+4 exp(-4.pi..sup.2Dt/L.sup.2)}
i.sub.ss=2 FADC.sub.o/L
[0035] where
[0036] i.sub.ss is the steady-state current following application
of the second electric potential;
[0037] i is the measured current which is a function of time
[0038] D is the diffusion coefficient of the cell, where this
coefficient may be determined from Fick's first law, i.e.
J(x,t)=-D.sup.dC(x,t)/.sub.- dx
[0039] L is the spacer thickness;
[0040] t is the time for the application of the 2.sup.nd electric
potential where t=0 for the beginning of the pulse
[0041] C.sub.0 is the preliminary concentration of the analyte;
[0042] F is faraday's constant, i.e. 9.648533 10.sup.4C/mol;
and
[0043] A is the area of the working electrode.
[0044] Using the above equations and steps, the observed first and
second time-current transients are used to determine the variable
.gamma. of the electrochemical cell employed in the subject method
and the preliminary concentration value of the analyte of interest
in the assayed physiological sample.
[0045] From the determined variable .gamma. and preliminary analyte
concentration value, a hematocrit correction factor is determined,
which hematocrit correction factor is used to obtain a hematocrit
corrected analyte concentration value from the initial or
preliminary analyte concentration value described above. The
hematocrit correction factor is a factor with which the preliminary
analyte concentration (typically less a background value) may be
multiplied in order to obtain a hematocrit corrected analyte
concentration value, i.e. a concentration value from which the
hematocrit component has been removed. The hematocrit correction
factor is a function of both the preliminary analyte concentration
value and the variable .gamma. of the electrochemical cell.
[0046] Any hematocrit correction factor that can be multiplied by
the preliminary concentration value (usually less a background
value, as described in greater detail below) may be employed in the
subject methods. One class of hematocrit correction factors that
find use in the subject methods are those that are derived from a
three dimensional graph of C.sub.0, .gamma. and .alpha.(C.sub.0,
.gamma.) obtained from experimental data using a wide range of
analyte and hematocrit values. The hematocrit correction factor
(.alpha.(C.sub.0, .gamma.)) is determined using the formula:
.alpha.(C.sub.0, .gamma.)=actual concentration/(C.sub.0-Background
Value)
[0047] (For example, where the analyte is glucose, .alpha.(C.sub.0,
.gamma.) in many embodiments equals the glucose concentration as
determined using the Yellow Springs Instrument glucose analyzer
model 23A (as described in U.S. Pat. No. 5,968,760 the disclosure
of which is herein incorporated by reference) divided by the
C.sub.0 less a background value, e.g. 22 mg/dL). This class of
hematocrit correction factors are typically equations which fit a
smooth surface function that minimizes the error between the
predicted and actual data. See e.g. the experimental section,
infra. One representative hematocrit correction factor that finds
use in the subject methods is:
1/((0.6637)+((4.9466*ln(C.sub.0))/C.sub.0)+(-0.4012*ln(.gamma.)))
[0048] In determining the hematocrit corrected concentration of
analyte according to the subject invention, the preliminary analyte
concentration (C.sub.0) as determined above, less a background
signal value, is multiplied by the hematocrit correction factor.
The background value that is subtracted from the preliminary
concentration value depends on the analyte being measured. For
glucose, this value typically ranges from about 0 to 40 mg/dL,
usually from about 8 to 25 mg/dL, where in many embodiments the
background value is about 22 mg/dL or is 22 mg/dL.
[0049] Generally, the following formula is employed to determine
the hematocrit corrected analyte concentration according to the
subject invention:
hematocrit corrected concentration hematocrit correction
factor.times.[C.sub.0-.beta.]
[0050] where
[0051] .beta. is the background value; and
[0052] C.sub.0 is the preliminary analyte concentration.
[0053] The above described methods yield a hematocrit corrected
analyte concentration value, i.e. a concentration value in which
the hematocrit component has been deconvoluted and removed. As
such, the above described methods provide for an accurate value of
the concentration of the analyte in the sample being assayed.
[0054] The above computational steps of the subject method may be
accomplished manually or through the use of an automated computing
means, where in many embodiments the use of an automated computing
means, such as is described in connection with the subject devices
discussed below, is of interest.
[0055] Devices
[0056] Also provided by the subject invention are meters for use in
practicing the subject invention. The subject meters are typically
meters for amperometrically measuring the hematocrit corrected
concentration of an analyte in a physiological sample. The subject
meters typically include: (a) a means for applying a first electric
potential to an electrochemical cell into which the sample has been
introduced and measuring cell current as a function of time to
obtain a first time-current transient; (b) a means for applying a
second electric potential to the electrochemical cell and measuring
cell current as a function of time to obtain a second time-current
transient; (c) a means for determining a preliminary analyte
concentration value and a variable .gamma. from said first and
second time-currents; and (d) a means for removing the hematocrit
component from the preliminary concentration value to derive the
hematocrit corrected analyte concentration in said sample. Means
(a) and (b) may be any suitable means, where representative means
are described in WO 97/18465 and U.S. Pat. No. 5,942,102; the
disclosures of which are herein incorporated by reference. Means
(c) and (d) are typically computing means present in the meter
which are capable of using the measured first and second time
current transients to ultimately obtain the hematocrit corrected
analyte concentration. As such, means (c) is typically a means that
is capable of determining the preliminary concentration of the
analyte of interest and the variable .gamma. from the first and
second time-current transients using the equations described above.
Likewise, means (d) is typically a means that is capable of
determining the hematocrit corrected analyte concentration using
the equations described above, where this means typically comprises
the hematocrit correction factor.
[0057] The following examples are offered by way of illustration
and not by way of limitation.
EXPERIMENTAL
[0058] I. Electrochemical Test Strip Preparation
[0059] An electrochemical test strip consisting of two metallized
electrodes oriented in a sandwich configuration was prepared as
follows. The top layer of the test strip was a gold sputtered Mylar
strip. The middle layer was a double-sided adhesive with a punched
hole that defined the reaction zone or area. The punched hole was a
circle with two juxtaposed rectangular inlet and outlet channels.
The bottom layer of the test strip was sputtered palladium on
Mylar. A film of ferricyanide and glucose dehydrogenase PQQ was
deposited on the palladium sputtered surface.
[0060] II. Generation of Experimental Data
[0061] First and second time current transients for a number of
different samples varying by glucose concentration and hematocrit
were obtained as follows. Sample was applied to the strip which
actuated an applied potential of -0.03V for a period of 10 seconds
which was then followed by a second pulse of +0.3V for a period of
3 to 10 seconds (where these electrode potentials are with respect
to the gold electrode).
[0062] III. Derivation of Hematocrit Correction Factor for
Glucose
[0063] For a wide range of glucose and hematocrit values measured
as described above, C.sub.0, the variable .gamma. and
.alpha.(C.sub.0, .gamma.) were derived.
[0064] C.sub.0 was derived using the equations:
i(t)=i.sub.ss{1+4 exp(-4.pi..sup.2Dt/L.sup.2)}
i.sub.ss=2 FADC.sub.0/L
[0065] where
[0066] i.sub.ss is the steady-state current following application
of the second electric potential;
[0067] i is the measured current which is a function of time
[0068] D is the diffusion coefficient of the cell, where this
coefficient may be determined from Fick's first law, i.e.
J(x,t)=-D.sup.dC(x,t)/.sub.- dx
[0069] L is the spacer thickness;
[0070] t is the time for the application of the 2.sup.nd electric
potential where t=0 for the beginning of the pulse;
[0071] C.sub.0 is the preliminary concentration of the analyte;
[0072] F is faraday's constant, i.e. 9.6485.times.10.sup.4C/mol;
and
[0073] A is the area of the electrode surface.
[0074] The variable .gamma. was derived using the equation:
.gamma.=i.sub.ss/i.sub.pp
[0075] where:
[0076] i.sub.ss is the steady-state current of the second applied
electric potential or second pulse; and
[0077] i.sub.pp is the average current from 8.5 to 9.5 seconds of
the 10 s long period during which the first pulse was applied.
[0078] .alpha.(C.sub.0, .gamma.) was determined using the
equation:
.alpha.(C.sub.0, .gamma.)=YSI concentration/(C.sub.0-22 mg/dL)
[0079] where YSI is the glucose concentration as determined using
the Yellow Springs Instrument glucose analyzer model 23A (as
described in U.S. Pat. No. 5,968,760 the disclosure of which is
herein incorporated by reference).
[0080] A three-dimensional graph of C.sub.0, .gamma. and
.alpha.(C.sub.0, .gamma.) as determined above for a wide range of
glucose and hematocrit values was prepared and is shown in FIG. 1.
A simple equation fit was then performed on the graph to define the
surface. The residual of the fitted data was monitored to ascertain
the quality of the model equation. The empirical equation was found
to be:
Hematocrit Correction
Factor=1/((0.6637)+((4.9466*ln(C.sub.0))/C.sub.0)+(--
0.4012*ln(.gamma.)))
[0081] The above correction factor was found to be valid for those
situations where the .gamma.>0.7 and C.sub.0>40 mg/dL.
[0082] IV. Comparison of Hematocrit Corrected Values to YSI
determined Values.
[0083] A prediction data set was generated by testing several
glucose strips with a wide range of glucose and hematocrit levels.
From this data a hematocrit correction equation was derived using a
model which fits the terms C.sub.0, .gamma., and .alpha.(C.sub.0,
.gamma.). It was found that using the hematocrit correction
equation on the prediction data set causes the majority of data
points to fall within +/-15%. It was also found that the bias of
the glucose results to 42% hematocrit, indicating that the
hematocrit effect on this data set is minimal. In order to confirm
this algorithm, another batch of glucose sesnsors was tested with a
different blood donor. It was found that the algorithm still
corrects for the hematocrit effect in a manner analogous to the
earlier findings.
[0084] The above results and discussion demonstrate that subject
invention provides a simple and powerful tool to obtain analyte
concentration values in which hematocrit derived error is
substantially if not entirely eliminated. As the subject methods
rely solely on the measurement of time-current transients, they may
be practiced with relatively simple electrochemical devices.
Furthermore, only small sample volumes need be employed and
relatively quick assay times are provided. As such, the subject
invention represents a significant contribution to the art.
[0085] All publications and patents cited in this specification are
herein incorporated by reference as if each individual publication
or patent were specifically and individually indicated to be
incorporated by reference. The citation of any publication is for
its disclosure prior to the filing date and should not be construed
as an admission that the present invention is not entitled to
antedate such publication by virtue of prior invention.
[0086] Although the foregoing invention has been described in some
detail by way of illustration and example for purposes of clarity
of understanding, it is readily apparent to those of ordinary skill
in the art in light of the teachings of this invention that certain
changes and modifications may be made thereto without departing
from the spirit or scope of the appended claims.
* * * * *