U.S. patent application number 12/543713 was filed with the patent office on 2009-12-31 for method of measuring blood component and sensor used in the method.
This patent application is currently assigned to PANASONIC CORPORATION. Invention is credited to Shin IKEDA, Teppei SHINNO.
Application Number | 20090321281 12/543713 |
Document ID | / |
Family ID | 34538287 |
Filed Date | 2009-12-31 |
United States Patent
Application |
20090321281 |
Kind Code |
A1 |
SHINNO; Teppei ; et
al. |
December 31, 2009 |
METHOD OF MEASURING BLOOD COMPONENT AND SENSOR USED IN THE
METHOD
Abstract
A sensor for blood component analysis that can correct the
effect of a hematocrit easily is provided. The sensor includes an
analysis portion including a working electrode, a counter
electrode, and a reagent portion. The reagent portion includes an
oxidoreductase that reacts with the blood component and a mediator,
and the blood component is measured by causing a redox reaction
between the blood component and the oxidoreductase in the presence
of the mediator and detecting a redox current generated by the
redox reaction by the working electrode and the counter electrode.
In this sensor, the reagent portion further includes a hemolyzing
agent (e.g., sodium cholate) for hemolyzing an erythrocyte, and
when detecting the redox current, the erythrocyte is hemolyzed with
the hemolyzing agent so as to cause hemoglobin released to an
outside of the erythrocyte to react with the mediator and a current
generated by this reaction also is detected to correct an effect of
a hematocrit.
Inventors: |
SHINNO; Teppei;
(Matsuyama-shi, JP) ; IKEDA; Shin; (Osaka,
JP) |
Correspondence
Address: |
Hamre, Schumann, Mueller & Larson, P.C.
P.O. Box 2902
Minneapolis
MN
55402-0902
US
|
Assignee: |
PANASONIC CORPORATION
Osaka
JP
|
Family ID: |
34538287 |
Appl. No.: |
12/543713 |
Filed: |
August 19, 2009 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
10954020 |
Sep 28, 2004 |
|
|
|
12543713 |
|
|
|
|
Current U.S.
Class: |
205/792 ;
204/403.01 |
Current CPC
Class: |
G01N 33/721 20130101;
C12Q 1/001 20130101; C12Q 1/006 20130101; G01N 33/66 20130101; G01N
27/3274 20130101 |
Class at
Publication: |
205/792 ;
204/403.01 |
International
Class: |
G01N 33/49 20060101
G01N033/49; G01N 27/26 20060101 G01N027/26 |
Foreign Application Data
Date |
Code |
Application Number |
Oct 2, 2003 |
JP |
2003-344761 |
Claims
1-20. (canceled)
21. A sensor for measuring a blood component, comprising: a reagent
portion; a channel for leading blood; and an air vent hole, wherein
blood is led to the reagent portion through the channel for leading
blood by capillary action due to the air vent hole, and an error
that counterbalances an error of a material that has an effect on
the sensor is obtained from the material, thereby counterbalancing
the error having the effect of the sensor.
22. The sensor according to claim 21, wherein the error that has
the effect on the sensor and the error that counterbalances the
error having the effect are obtained with respect to an
amperometric response.
23. The sensor according to claim 21, wherein the error that has
the effect on the sensor is a negative error, and the error that
counterbalances the error having the effect is a positive
error.
24. A method that uses a sensor for measuring a blood component,
comprising: a reagent portion; a channel for leading blood; and an
air vent hole; the method comprising: leading blood to the reagent
portion through the channel for leading blood by capillary action
due to the air vent hole; and obtaining an error that
counterbalances an error of a material that has an effect on the
sensor from the material, thereby counterbalancing the error having
the effect on the sensor.
25. The method according to claim 24, wherein the error that has
the effect on the sensor and the error that counterbalances the
error having the effect are obtained with respect to the
amperometric response.
26. The sensor according to claim 24, wherein the error that has
the effect on the sensor is a negative error, and the error that
counterbalances the error having the effect is a positive error.
Description
CROSS-REFERENCE TO RELATED APPLICATION
[0001] This application is a Continuation of application Ser. No.
10/954,020, filed Sep. 28, 2004, which application is incorporated
herein by reference.
BACKGROUND OF THE INVENTION
[0002] 1. Field of the Invention
[0003] The present invention relates to a method of measuring a
blood component and a sensor used in the method.
[0004] 2. Related Background Art
[0005] Conventionally, sensors for blood component measurement have
been used for clinical test, self-measurement of blood glucose
level by diabetics, etc. The configuration of the sensor for blood
component measurement is such that, for example, a cover is
disposed on an insulating substrate having a working electrode and
a counter electrode on its surface, with a spacer intervening
between the cover and the insulating substrate. On the working
electrode and the counter electrode, a reagent containing an
oxidoreductase, a mediator (an electron carrier), and the like is
provided, thereby forming an analysis portion. The analysis portion
communicates with one end of a channel for leading blood to the
analysis portion. The other end of the channel is open toward the
outside of the sensor so as to serve as a blood supply port. Blood
component analysis (e.g., analysis of blood glucose level) using
the sensor configured as above is carried out in the following
manner, for example. First, the sensor is set in a dedicated
measuring device (a meter). Then, a fingertip or the like is
injured with a lancet to cause bleeding, and the blood supply port
of the sensor is brought into contact with the blood that has come
out. The blood is drawn into the channel of the sensor by capillary
action and flows through the channel to be led to the analysis
portion where the blood comes into contact with the reagent. Then,
a redox reaction occurs between a blood component and the
oxidoreductase so that a current flows via the mediator. The
working electrode and the counter electrode detect the current, and
the measuring device converts the detected current into an amount
of the blood component and displays the value obtained by the
conversion.
[0006] In the above-described manner, the sensor can measure the
blood component. However, since the obtained measured value might
be affected by a hematocrit (Hct), it might be necessary to remove
the effect of Hct in order to obtain an accurate measured value.
One example of a method of removing the effect of Hct is preparing
a correction table beforehand using a sample with a known Hct and
then correcting the measured value using this correction table (see
JP 11(1999)-194108 A, for example). Another example is correcting a
Hct using a parameter that has been set beforehand (see WO
02/44705, for example). However, these methods require a laborious
correction process such as providing a correction table beforehand
or performing a complicated calculation using a parameter.
SUMMARY OF THE INVENTION
[0007] The present invention was made in light of the foregoing
problems, and it is an object of the present invention to provide a
method and a sensor that can measure a blood component without a
laborious correction process.
[0008] In order to achieve the above object, the present invention
provides a method of measuring a blood component, including:
causing a redox reaction between the blood component and an
oxidoreductase in the presence of a mediator; detecting a redox
current generated by the redox reaction by electrodes; and
converting the detected current value into an amount of the blood
component, wherein when detecting the redox current, an erythrocyte
is hemolyzed so as to cause hemoglobin released to an outside of
the erythrocyte to react with the mediator and a current generated
by this reaction also is detected to correct an effect of a
hematocrit.
[0009] The present invention also provides a sensor for measuring a
blood component, including an analysis portion, the analysis
portion including: a working electrode; a counter electrode; and a
reagent portion. The reagent portion includes an oxidoreductase
that reacts with the blood component and a mediator, and the blood
component is measured by causing a redox reaction between the blood
component and the oxidoreductase in the presence of the mediator
and detecting a redox current generated by the redox reaction by
the working electrode and the counter electrode. In this sensor,
the reagent portion further includes a hemolyzing agent for
hemolyzing an erythrocyte, and when detecting the redox current,
the erythrocyte is hemolyzed with the hemolyzing agent so as to
cause hemoglobin released to an outside of the erythrocyte to react
with the mediator and a current generated by this reaction also is
detected to correct an effect of a hematocrit.
[0010] Note here that a greater Hct means a greater amount of
hemoglobin. That means, when the erythrocyte is hemolyzed to cause
the hemoglobin released to the outside of the erythrocyte to react
with the mediator, the current generated by this reaction also is
greater. Therefore, by detecting this current along with the
current generated by the redox reaction of the blood component,
even when the current generated by the redox reaction is smaller
than the actual value due to the effect of the Hct, the current
value that has been corrected to remove the effect of the Hct can
be obtained by the electrodes. Thus, according to the present
invention, by electrochemically detecting the blood component and
also the hemoglobin that varies depending on Hct, the effect of Hct
can be corrected automatically by performing current detection only
once. Therefore, a complicated correction process is not
necessary.
BRIEF DESCRIPTION OF THR DRAWINGS
[0011] FIG. 1A is an exploded perspective view showing an example
of a sensor according to the present invention, and FIG. 1B is a
cross-sectional view of the same.
[0012] FIG. 2 is a view illustrating the principle of the present
invention.
[0013] FIG. 3 is a graph showing the change in measured current
with the change in Hct in an example of the present invention.
[0014] FIG. 4 is a graph showing the change in measured current
with the change in Hct in a comparative example of the present
invention.
[0015] FIG. 5 is a graph showing the change in measured current
with the change in Hct in another example of the present
invention.
[0016] FIG. 6 is a graph showing the change in measured current
with the change in Hct in the case where sodium taurocholate is
added to a reagent solution in still another example of the present
invention.
[0017] FIG. 7 is a graph showing the change in measured current
with the change in Hct in the case where sodium taurodeoxycholate
is added to a reagent solution in still another example of the
present invention.
[0018] FIG. 8 is a graph showing the change in measured current
with the change in Hct in the case where sodium glycocholate is
added to a reagent solution in still another example of the present
invention.
DETAILED DESCRIPTION OF THE INVENTION
[0019] Hereinafter, the present invention will be described in
detail.
[0020] In the measurement method of the present invention, the
hemolysis preferably is caused by a membrane protein solubilizer so
as to allow the hemolysis to be caused in a simple manner without
performing a laborious operation. However, in the present
invention, means for causing the hemolysis is not limited thereto,
and can be, for example, physical means such as an osmotic shock
procedure using an anisotonic solution, an ultrasonic treatment, or
a freezing and thawing method that causes hemolysis by repeating
freezing and thawing.
[0021] In the measurement method and the sensor of the present
invention, the membrane protein solubilizer is not particularly
limited as long as it can hemolyze erythrocytes. Note here that the
term "hemolysis" as used herein refers to a phenomenon in which a
membrane of an erythrocyte is broken and hemoglobin and the like
contained in the erythrocyte are release to the outside of the
erythrocyte. Examples of the membrane protein solubilizer include
lipase, saponins, lysozyme, inorganic salts, and detergents. Among
them, detergents are more preferable. Examples of the detergents
include ionic detergents such as anionic detergents, cationic
detergents, and amphoteric detergents, nonionic detergents, and
cholic acid-based detergents. Among them, cholic acid-based
detergents are preferable in terms of simplicity in preparing a
reagent and the crystal condition of a reagent portion. Examples of
the cholic acid-based detergent include cholic acid, sodium
cholate, cholic acid methyl ester, chenodeoxycholic acid, sodium
chenodeoxycholate, diphenylglycolic acid (benzilic acid),
deoxycholic acid, sodium deoxycholate, sodium
glycochenodeoxycholate, glycocholic acid, sodium glycocholate,
glycodeoxycholic acid, sodium glycodeoxycholate, glycolic acid,
sodium glycolate, sodium glycolithocholate, lithocholic acid,
sodium thioglycolate, sodium taurocholate, sodium
taurodeoxycholate, sodium tauroursodeoxycholate, sodium
ursodeoxycholate, and ursodeoxycholic acid. They may be used
individually or two or more of them may be used together. Among the
above-described cholic acid-based detergents, sodium cholate,
sodium deoxycholate, sodium glycocholate, sodium glycodeoxycholate,
sodium taurocholate, and sodium taurodeoxycholate are preferable,
and sodium cholate, sodium glycocholate, sodium taurocholate, and
sodium taurodeoxycholate are particularly preferable. In addition
to the above-described detergents, the following detergents also
can be used, for example: sodium lauryl sulfate (SDS);
N,N-bis(3-D-gluconamidopropyl)cholamide (BIGCHAP);
3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS);
N,N-bis(3-D-gluconamidopropyl)deoxycholamide (deoxy-BIGCHAP);
3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonate
(CHAPSO); n-decanoyl-N-methylglucamide (MEGA-10);
n-nonanoyl-N-methylglucamide (MEGA-9); n-octanoyl-N-methylglucamide
(MEGA-8); n-octyl-.beta.-D-thioglucoside;
n-octyl-.beta.-D-maltoside; n-octyl-.beta.-D-glucoside; sucrose
monolaurate (SM1200); sucrose monocaprate (SM1000); and sucrose
monocholate.
[0022] In the measurement method and the sensor of the present
invention, the amount of the membrane protein solubilizer is not
particularly limited, but may be, for example, 0.01 mM to 100 mM,
preferably 0.1 mM to 50 mM, and particularly preferably 0.2 mM to
2.0 mM per one measurement or one sensor.
[0023] In the measurement method and the sensor of the present
invention, the mediator is not particularly limited. Examples of
the mediator include potassium ferricyanide, p-benzoquinone,
p-benzoquinone derivatives, phenazine methosulfate, methylene blue,
ferrocene, and ferrocene derivatives. Among them, potassium
ferricyanide is preferable. The amount of the mediator is not
particularly limited, but may be, for example, 0.1 mM to 1000 mM,
preferably 1 mM to 500 mM, and more preferably 5 mM to 200 mM per
one sensor or one measurement.
[0024] In the measurement method and the sensor of the present
invention, an analyte is not particularly limited as long as it is
a blood component, and may be, for instance, glucose, lactic acid,
uric acid, bilirubin, cholesterol, or the like. The oxidoreductase
may be an oxidoreductase that reacts with a blood component as an
analyte, and examples thereof include glucose oxidase, lactate
oxidase, cholesterol oxidase, bilirubin oxidase, glucose
dehydrogenase, and lactate dehydrogenase. The amount of the
oxidoreductase may be, for example, 0.1 U to 100 U, preferably 0.5
U to 50 U, and more preferably 1 U to 10 Upper one sensor or one
measurement.
[0025] In the sensor of the present invention, it is preferable
that the reagent portion further contains a hydrophilic polymer, an
enzyme stabilizer, and a crystal homogenizing agent.
[0026] The hydrophilic polymer serves to impart viscosity to a
reagent solution so that, when preparing the reagent portion by
drying the reagent solution, a homogenous reagent portion is formed
on the electrodes easily and the adhesion between the electrode and
the reagent portion is enhanced. The hydrophilic polymer also
serves to improve the crystal condition of the reagent portion
after being dried. Examples of the hydrophilic polymer include
carboxymethyl cellulose (CMC), hydroxyethyl cellulose,
hydroxypropyl cellulose, methyl cellulose, ethylcellulose, ethyl
hydroxyethyl cellulose, carboxyethyl cellulose, polyvinyl alcohol,
polyvinylpyrrolidone, polyamino acid such as polylysine,
polystyrene sulfonate, gelatin and derivatives thereof, polyacrylic
acid and salts thereof, polymethacrylic acid and salts thereof,
starch and derivatives thereof, maleic anhydride polymer and salts
thereof, and agarose gel and derivatives thereof. They may be used
individually or two or more of them may be used together. Among
them, CMC is preferable. The ratio of the hydrophilic polymer to
the entire reagent solution for preparing a reagent portion may be,
for example, 0.001 wt % to 5 wt %, preferably 0.005 wt % to 2.5 wt
%, and more preferably 0.01 wt % to 1.0 wt %.
[0027] As the enzyme stabilizer, sugar alcohol may be used.
Examples of the sugar alcohol include chain polyhydric alcohols and
cyclic sugar alcohols, such as sorbitol, maltitol, xylitol,
mannitol, lactitol, reduced paratinose, arabinitol, glycerol,
ribitol, galactitol, sedoheptitol, perseitol, volemitol,
styracitol, polygalitol, iditol, talitol, allitol, inositol,
hydrogenated glucose syrup, and isylitol. Note here that
stereoisomers, substitution products, and derivatives of these
sugar alcohols may also be used as the enzyme stabilizer. These
sugar alcohols may be used individually or two or more of them may
be used together. Among them, maltitol is preferable. The amount of
the enzyme stabilizer may be, for example, 0.01 mM to 500 mM,
preferably 0.05 mM to 100 mM, and more preferably 0.1 mM to 50 mM
per one measurement or one sensor.
[0028] The crystal homogenizing agent serves to homogenize the
crystal condition of the reagent portion. As the crystal
homogenizing agent, an amino acid may be used, for example.
Examples of the amino acid include glycine, alanine, valine,
leucine, isoleucine, serine, threonine, methionine, asparagine,
glutamine, arginine, lysine, histidine, phenylalanine, tryptophan,
proline, sarcosine, betaine, taurine, and salts, substitution
products, and derivatives of these amino acids. They may be used
individually or two or more of them may be used together. Among
them, glycine, serine, proline, threonine, lysine, and taurine are
preferable, and taurine is more preferable. The amount of the
crystal homogenizing agent may be, for example, 0.1 mM to 1000 mM,
preferably 5 mM to 500 mM, and more preferably 10 mM to 300 mM per
one measurement or one sensor.
[0029] Next, the configuration of the sensor of the present
invention will be described. For example, in the sensor of the
present invention, a working electrode and a counter electrode are
disposed on an insulating substrate, thereby forming an analysis
portion. A reagent portion further is disposed on the analysis
portion. The analysis portion communicates with one end of a
channel for leading blood to the analysis portion, and the other
end of the channel is open toward the outside of the sensor,
thereby allowing this opening to serve as a blood supply port. On
the insulating substrate, a cover is disposed with a spacer
intervening therebetween. Preferably, the sensor further includes a
detecting electrode that is located farther from the blood supply
port than the analysis portion so that whether or not blood is
supplied to the analysis portion is detected by this detecting
electrode.
[0030] FIG. 1 shows one example of the sensor of the present
invention configured as above. FIG. 1A is an exploded perspective
view of the sensor, and FIG. 1B is a cross-sectional view of the
same. As shown in FIG. 1, in this sensor, a working electrode 14
and a counter electrode 15 are formed on an insulating substrate
11, and a reagent portion 19 is disposed on these electrodes,
thereby forming an analysis portion. On the insulating substrate
11, a detecting electrode 16 further is formed, which is located
farther from the blood inlet port side than the working electrode
14 and the counter electrode 15. The reagent portion 19 contains
the oxidoreductase such as glucose oxidase as described above, the
mediator as described above, a hemolyzing agent such as cholic
acid, the hydrophilic polymer as described above, the enzyme
stabilizer as described above, the crystal homogenizing agent as
described above, and the like. The type and the blend ratio of
these reagents are as described above. A cover 13 is disposed on
the insulating substrate 11 so as to cover an entire area excluding
one end portion (the end portion on the right in FIG. 1) with a
spacer 12 intervening therebetween. The analysis portion
communicates with a channel 17 for leading blood to the analysis
portion. The channel 17 extends to the other end portion (the end
portion on the left in FIG. 1) of the sensor, and the tip of the
channel 17 on the other end portion side is open toward the outside
of the sensor so as to serve as a blood inlet port. The working
electrode 14, the counter electrode 15, and the detecting electrode
16 are connected to leads, respectively. These leads extend to the
above-described one end portion of the sensor with the tip of each
lead not being covered with the cover but being exposed. The cover
13 has an air vent hole 18 for enhancing the capillary action at a
portion corresponding to the rear side of the channel 17.
[0031] In the present invention, the material for the insulating
substrate is not particularly limited, and may be, for example,
polyethylene terephthalate (PET), polycarbonate (PC), polyimide
(PI), polyethylene (PE), polypropylene (PP), polystyrene (PS),
polyvinyl chloride (PVC), polyoxymethylene (POM), monomer-cast
nylon (MC), polybutylene terephthalate (PBT), polymethyl
methacrylate (PMMA), an ABS resin (ABS), or glass. Among them,
polyethylene terephthalate (PET), polycarbonate (PC), and polyimide
(PI) are preferable, and polyethylene terephthalate (PET) is more
preferable. The size of the insulating substrate is not
particularly limited. For example, the insulating substrate may
have an overall length of 5 mm to 100 mm, a width of 3 mm to 50 mm,
and a thickness of 0.1 mm to 2 mm; preferably an overall length of
10 mm to 50 mm, a width of 3 mm to 20 mm, and a thickness of 0.2 mm
to 1 mm; and more preferably an overall length of 15 mm to 30 mm, a
width of 5 mm to 10 mm, and a thickness of 0.3 mm to 0.6 mm.
[0032] The electrodes and leads on the insulating substrate may be
formed, for example, by forming a conductive layer with gold,
platinum, palladium, or the like by sputtering or vapor deposition
and then processing the conductive layer into a particular
electrode pattern with a laser. Examples of the laser include YAG
lasers, CO.sub.2 lasers, and excimer lasers.
[0033] The reagent portion can be formed, for example, by
dissolving a predetermined reagent in water or a buffer solution
and then drying it. For example, in a 0.01 wt % to 2.0 wt % CMC
aqueous solution, 0.1 U/sensor to 5.5 U/sensor of PQQ-GDH, 10 mM to
200 mM of potassium ferricyanide, 0.05 mM to 30 mM of maltitol, 10
mM to 300 mM of taurine, and 0.02 mM to 5.0 mM of sodium cholate
are added and dissolved. The reagent portion can be formed by
dropping the thus-obtained solution on the analysis portion (on the
working electrode and the counter electrode) of the substrate and
then drying it. The drying may be either air drying or forced
drying using warm air. However, if the temperature of the warm air
is too high, there is a possibility that the enzyme contained in
the solution might be deactivated. Thus, the temperature of the
warm air preferably is around 50.degree. C.
[0034] In the present invention, the material for the spacer is not
particularly limited. For example, the same material as that for
the insulating substrate can be used. The size of the spacer also
is not particularly limited. For example, the spacer may have an
overall length of 5 mm to 100 mm, a width of 3 mm to 50 mm, and a
thickness of 0.01 mm to 1 mm; preferably an overall length of 10 mm
to 50 mm, a width of 3 mm to 20 mm, and a thickness 0.05 mm to 0.5
mm; and more preferably an overall length of 15 mm to 30 mm, a
width of 5 mm to 10 mm, and a thickness of 0.05 mm to 0.25 mm. The
spacer has a cut-away portion that serves as a channel for leading
blood. The cut-away portion may have, for example, an overall
length of 1 mm to 30 mm and a width of 0.05 mm to 10 mm, preferably
an overall length of 2 mm to 10 mm and a width of 0.3 mm to 5 mm,
and more preferably an overall length of 2 mm to 10 mm and a width
of 0.5 mm to 2 mm. The cut-away portion may be formed, for
instance, by using a laser, a drill, or the like, or by forming the
spacer using a die that can form the spacer provided with the
cut-away portion.
[0035] In the present invention, the material for the cover is not
particularly limited. For example, the same material as that for
the insulating substrate can be used. It is more preferable that a
portion of the cover corresponding to the ceiling of the sample
supply channel is subject to a treatment for imparting
hydrophilicity. The treatment for imparting hydrophilicity may be
carried out by, for example, applying a detergent or introducing a
hydrophilic functional group such as a hydroxyl group, a carbonyl
group, or a carboxyl group to the cover surface by plasma
processing or the like. The size of the cover is not particularly
limited. For example, the cover may have an overall length of 5 mm
to 100 mm, a width of 3 mm to 50 mm, and a thickness of 0.01 mm to
0.5 mm; preferably an overall length of 10 mm to 50 mm, a width of
3 mm to 20 mm, and a thickness of 0.05 mm to 0.25 mm; and more
preferably an overall length of 15 mm to 30 mm, a width of 5 mm to
10 mm, and a thickness of 0.05 mm to 0.1 mm. The cover preferably
has an air vent hole, which may have, for example, a maximum
diameter of 0.01 mm to 10 mm, preferably 0.05 mm to 5 mm, and more
preferably 0.1 mm to 2 mm. The air vent hole may be formed, for
instance, by perforating the cover with a laser, a drill, or the
like, or by forming the cover using a die that can form the cover
provided with the air vent hole.
[0036] This sensor can be produced by laminating the insulating
substrate, the spacer, and the cover in this order and integrating
them. The integration can be achieved by adhering these three
components with an adhesive or through heat-sealing. As the
adhesive, an epoxy adhesive, an acrylic adhesive, a polyurethane
adhesive, a thermosetting adhesive (a hot melt adhesive or the
like), a UV curable adhesive, or the like can be used, for
example.
[0037] Measurement of blood glucose level using this sensor can be
carried out in the following manner, for example. First, a
fingertip or the like is punctured with a dedicated lancet to cause
bleeding. On the other hand, the sensor is set in a dedicated
measuring device (a meter). The blood inlet port of the sensor that
is set in the measuring device is brought into contact with the
blood that has come out, so that the blood is led to the analysis
portion of the sensor by capillary action. In the analysis portion,
glucose in the blood reacts with the oxidoreductase such as glucose
oxidase contained in the reagent. On the other hand, after a lapse
of a certain period after the detecting electrode detects the
supply of the blood to the analysis portion, a constant voltage is
applied between the working electrode and the counter electrode. As
a result, a redox current flows. At this time, erythrocytes
contained in the blood have been hemolyzed by the hemolyzing agent
in the reagent portion 19, thereby releasing hemoglobin to the
outside of the erythrocytes. The hemoglobin released to the outside
reacts with the mediator, and a current generated by this reaction
is detected by the electrodes simultaneously with the redox
current. The detected current is measured by the measuring device,
which converts the measured value into a glucose concentration and
displays the value obtained by the conversion.
[0038] In the measurement using this sensor, the effect of Hct is
corrected automatically. The reason for this will be described with
reference to FIG. 2. As shown in the upper left graph of FIG. 2, an
amount of hemoglobin in blood increases in keeping with Hct.
Accordingly, an amount of reduced mediator generated by the
electron exchange reaction between the hemoglobin and the mediator
also increases. Although a reduced mediator generated by an enzyme
reaction actually is to be measured, a reduced mediator also is
generated through the above reaction, which causes an amperometric
response obtained finally to become greater than it should be
(hereinafter this phenomenon is referred to as a "positive error").
On the other hand, it has been known that an increase in Hct, i.e.,
an increase in blood cell (solid) components considerably affects
the elementary processes (a nonuniform electron transfer reaction,
diffusion, etc.) of the electrode reaction by electrode active
species. Thus, an increase in Hct also causes the amperometric
response obtained finally to become smaller than it should be
(hereinafter this phenomenon is referred to as a "negative error").
In general, in a system without a membrane protein solubilizer, the
above-described negative error is remarkable because solubilization
of erythrocytes is not promoted in such a system. Thus, as shown in
the upper right graph of FIG. 2, the amperometric response tends to
decrease as Hct increases. On this account, by adding a membrane
protein solubilizer to a sensor system so as to promote the
solubilization of erythrocytes, it becomes possible to
counterbalance the positive error and the negative error. As a
result, it is possible to realize more accurate quantification of a
blood component with the Hct value dependency of a sensor response
being minimized (see the lower graph of FIG. 2).
[0039] Note here that the above sensor merely is an example of a
sensor according to the present invention, and a sensor without a
detecting electrode, for example, also falls within the scope of
the present invention.
Example 1
[0040] Hereinafter, examples of the present invention will be
described along with a comparative example.
[0041] Sensors having the configuration as shown in FIG. 1 were
produced in the manner described above. A reagent solution having
the following composition was prepared, which was dropped on an
analysis portion of each sensor and then dried to form a reagent
portion.
(Composition of Reagent Portion)
[0042] enzyme (PQQ-GDH)
[0043] mediator (potassium ferricyanide)
[0044] hydrophilic polymer (CMC)
[0045] enzyme stabilizer (maltitol)
[0046] crystal homogenizing agent (taurine)
[0047] membrane protein solubilizer (sodium cholate: 1.2 mM)
[0048] On the other hand, from two types of human whole blood with
glucose concentrations of 100 mg/dL and 400 mg/dL, six types of
human whole blood samples were prepared by adjusting the Hct to
25%, 45%, and 65%.
[0049] With regard to each sample, the measurement was carried out
in the following manner. The sensor was set in a dedicated
measuring device (a meter), and the blood inlet port of the sensor
was brought into contact with the sample so that the sample was led
to the analysis portion by capillary action. The measurement was
started when the sample was detected by the detecting electrode.
After a lapse of 3.5 seconds, a constant voltage of +0.2 V was
applied between the working electrode and the counter electrode,
and a current value after 1.5 seconds was measured. The number (n)
of times the measurement was performed was n=10 with regard to each
sample, and the average of the obtained measured values is shown in
the graph of FIG. 3. In the graph of FIG. 3, the detected current
with regard to each of the samples with the Hct of 45% was set as a
standard point, and the deviations (%) of the detected currents
with regard to the samples with the other Hct values from this
standard point are shown.
Comparative Example
[0050] Sensors were produced in the same manner as in Example 1
except that the hemolyzing agent was not used, and the measurement
of current using these sensors was carried out in the same manner
as in Example 1. The results are shown in the graph of FIG. 4. In
the graph of FIG. 4, the detected current with regard to each of
the samples with the Hct of 45% was set as a standard point, and
the deviations (%) of the detected currents with regard to the
samples with the other Hct values from this standard point are
shown, as in the graph of FIG. 3.
[0051] As can be seen from the graph of FIG. 3, the current values
obtained by the sensors according to Example 1 were substantially
constant even under the varying Hct. In contrast, as can be seen
from the graph of FIG. 4, the current values obtained by the
sensors according to the comparative example varied greatly with
the change in Hct.
Example 2
[0052] Sensors were produced in the same manner as in Example 1. In
these sensors, the reagent portion contained the same components as
those in Example 1, but the amount of sodium cholate as the
membrane protein solubilizer was changed. More specifically, in the
present example, three types of sensors, namely, the sensor with
0.8 mM of sodium cholate, the sensor with 1.8 mM of sodium cholate,
and the conventional sensor without the membrane protein
solubilizer were produced.
[0053] The measurement was carried out in the same manner as in
Example 1. The conditions for the current measurement and the
number (n) of times the measurement was performed also were the
same as those in Example 1. FIG. 5 shows the results of the
measurement performed with regard to three types of human whole
blood samples prepared by adjusting the Hct of human whole blood
with glucose concentration of 100 mg/dL to 25%, 45%, and 65%. In
the graph of FIG. 5, the detected current with regard to the sample
with the Hct of 45% was set as a standard point, and the deviations
(%) of the detected currents with regard to the samples with the
other Hct values from this standard point are shown.
[0054] As is clear from FIG. 5, the effect of Hct was reduced
gradually with an increase in the concentration of the sodium
cholate added to the reagent portion.
Example 3
[0055] Sensors were produced in the same manner as in Example 1. In
these sensors, the composition of the reagent portion was the same
as in Example 1 except that the type of the membrane protein
solubilizer was changed. More specifically, in the present example,
three types of sensors respectively employing the following
membrane protein solubilizers were produced.
(Membrane Protein Solubilizer)
[0056] sodium taurocholate (1.2 mM)
[0057] sodium taurodeoxycholate (1.2 mM)
[0058] sodium glycocholate (1.2 mM)
[0059] Using these sensors, the measurement was performed in the
same manner as in Example 1 with regard to six types of human whole
blood samples prepared by adjusting the Hct of two types of human
whole blood with glucose concentrations of 100 mg/dL and 400 mg/dL
to 25%, 45%, and 65%. FIG. 6 shows the result of the measurement
using the sensor employing sodium taurocholate as the membrane
protein solubilizer, FIG. 7 shows the result of the measurement
using the sensor employing sodium taurodeoxycholate as the membrane
protein solubilizer, and FIG. 8 shows the result of the measurement
using the sensor employing sodium glycocholate as the membrane
protein solubilizer. In the graphs of FIG. 6, FIG. 7, and FIG. 8,
the detected current with regard to each of the samples with the
Hct of 45% was set as a standard point, and the deviations (%) of
the detected currents with regard to the samples with the other Hct
values from this standard point are shown. In the present example,
the conditions for the current measurement and the number (n) of
times the measurement was performed also were the same as those in
Example 1.
[0060] As clear from FIG. 6, FIG. 7, and FIG. 8 in comparison with
FIG. 4 directed to a comparative example of Example 1, the membrane
protein solubilizers used in the present example also could reduce
the effect of Hct, as in the case of sodium cholate used in Example
1.
[0061] Although Examples 1, 2, and 3 are directed to a sensor for
measuring a glucose concentration in blood, it is to be noted that
an analyte or a measuring method is not limited thereto. For
example, an analyte may be lactic acid, cholesterol, uric acid, or
bilirubin. Moreover, although Examples 1, 2, and 3 are directed to
an example where a current was measured using a sensor with a
three-electrode structure including the working electrode 14, the
counter electrode 15, and the detecting electrode 16 as shown in
FIG. 1, it is to be noted that a sensor with a two-electrode
structure without a detecting electrode also is within the scope of
the present invention, and either of the three-electrode structure
or the two-electrode structure may be used in the present
invention. However, it is to be noted here that a sensor with three
electrodes can achieve more accurate measurement than a sensor with
two electrodes.
[0062] According to the measurement method and the sensor of the
present invention, the effect of Hct can be corrected automatically
and easily. Therefore, the measurement method and the sensor of the
present invention are useful in measurement of a blood
component.
[0063] Specific embodiments and examples described in the detailed
description of the present invention are intended merely to clarify
the technical details of the present invention. The present
invention should not be limited to such specific examples to be
understood narrowly. The present invention can be changed variously
to be carried out within the spirit of the present invention and
the range of the following claims.
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