JP2019035748A - Measurement method and measuring device - Google Patents

Abstract

To enable a measurement object component in a sample to be measured with high accuracy.SOLUTION: A measurement method of the present invention includes: applying a direct-current first signal having a first value to a sample in a flow channel and measuring a first electrical response value using, among a first electrode provided with an enzyme-containing reagent, a second and a third electrode not provided with the reagent, and a fourth electrode different from the first through third electrodes, the first electrode as an acting electrode and the fourth electrode as a counter electrode; applying a direct-current second signal having a second value higher than the first value to the sample in the flow channel continuously for a prescribed duration using the second electrode and the third electrode as an acting electrode and a counter electrode; measuring a second electrical response value to the second signal within a prescribed time that indicates the amount of electric charges generated by electrolysis of water content in the sample; and correcting the value obtained from the first electrical response value on the basis of the second electrical response value.SELECTED DRAWING: Figure 5

Description

  The present invention relates to a measurement method and a measurement apparatus.

  Conventionally, there is a technique of measuring a glucose concentration value (glucose value) and a hematocrit value (Hct value) in blood, which is an example of a biological sample, and correcting the glucose value with the Hct value. As a measuring method of the glucose value and the Hct value, for example, the Hct value is measured using a working electrode provided with a mediator and a counter electrode, and the glucose value is measured using a working electrode provided with a reagent and a counter electrode. There exists a method (for example, refer patent document 1).

  Alternatively, there is a method in which an Hct value is measured by applying an alternating voltage and a glucose value is measured by applying a direct voltage to a working electrode and a counter electrode each provided with a reagent (see, for example, Patent Document 2). Furthermore, there is a method in which a common working electrode is used for the measurement of the glucose value and the measurement of the Hct value and both measurements are performed by applying a DC voltage (for example, see Patent Document 3). Alternatively, the first electrical response to the first signal input to the first electrode pair that can contact the sample is measured, and the second electric signal input to the second electrode pair that can contact the sample. A second electrical response to the second signal, the value of which changes from a first level to a second level and then remains at the second level for a certain period of time. There is a method of measuring a peak value of a response signal with respect to a change and correcting a value indicating an amount of a measurement target component of the sample obtained from a first electrical response based on the peak value of the response signal (for example, a patent) Reference 4).

Japanese Patent No. 4611208 JP-T-2006-510124 JP 2005-147990 A JP 2014-232102 A

  An object of this invention is to provide the measuring method and measuring apparatus which enable the measurement of the measuring object component in a sample with high precision using a new method.

  One aspect of the present invention is a measurement method for measuring a measurement target component contained in a biological sample using an analysis tool in which a plurality of electrodes are arranged in a flow path. The measurement method includes a first electrode provided with a reagent containing an enzyme, a second electrode and a third electrode not provided with the reagent, the first electrode, and the second electrode, which are included in the plurality of electrodes. Of the electrode and the fourth electrode different from the third electrode, the first electrode is used as a working electrode and the fourth electrode is used as a counter electrode, and a first DC signal having a first value is generated in the flow path. Applying to the sample, measuring a first electrical response value of the sample with respect to the first signal, and using the second electrode and the third electrode as a working electrode and a counter electrode. A step of continuously applying a second DC signal having a second value higher than the value to the sample in the flow path for a predetermined time, and an amount of electric charge generated by electrolysis of moisture in the sample, A second electrical response value of the sample to a second signal And measuring within serial predetermined time, based on the second electrical response value, characterized in that it comprises a and a step of correcting the values obtained from the first electrical response value.

  Another aspect of the present invention is a measurement method for measuring a measurement target component contained in a biological sample using an analysis tool in which a plurality of electrodes are arranged in a flow path. This measurement method uses the first electrode of the plurality of electrodes, the first electrode provided with a reagent containing an enzyme, and the second electrode and the third electrode not provided with the reagent. Applying a DC first signal having a first value to the sample in the flow path using one of the second electrode and the third electrode as a counter electrode, and using the one of the second electrode and the third electrode as a counter electrode; A step of measuring a first electrical response value of the sample; and a second DC signal having a second value higher than the first value by using the second electrode and the third electrode as a working electrode and a counter electrode. A second electrical response value of the sample with respect to the second signal, which indicates a charge amount generated by electrolysis of moisture in the sample, continuously applied to the sample in the flow path for a predetermined time. Measuring within the predetermined time; and Based on the electrical response value, characterized in that it comprises a and a step of correcting the values obtained from the first electrical response value.

  Another aspect of the present invention is a measuring apparatus having the same characteristics as the measuring method described above.

  According to the present invention, it is possible to measure a measurement target component in a sample with high accuracy.

FIG. 1A is a top view of an analysis tool (a biosensor having four electrodes) according to the embodiment, and FIG. 1B is a side view of the biosensor shown in FIG. FIG. 2A is a top view of the analysis tool (biosensor having three electrodes) according to the embodiment, and FIG. 2B is a side view of the biosensor shown in FIG. 3A and 3B show a configuration example of the measuring apparatus according to the embodiment. FIG. 4 is a flowchart illustrating an example of a measurement method using the biosensor and the measurement device according to the embodiment. FIG. 5A shows a correspondence relationship (linearity of the Hct calibration curve) between the second electrical response value and the Hct value when the glucose value is constant. FIG. 5B shows a temporal change (time course) of the second electrical response value when the glucose value is constant, and FIG. 5C is a time of the first electrical response value when the glucose value is constant. Change. FIG. 6A shows the temporal change of the second electrical response value when the Hct value is constant, and FIG. 6B shows the temporal change of the first electrical response value when the Hct value is constant.

  Hereinafter, a measurement method and a measurement apparatus according to the embodiment will be described. In the embodiment, a measurement method for measuring a measurement target component contained in a biological sample using an analysis tool in which a plurality of electrodes are arranged in a flow path will be described. The measurement method includes a first measurement method and a second measurement method.

  The first measurement method includes a first electrode provided with a reagent containing an enzyme, a second electrode and a third electrode not provided with a reagent, a first electrode, a second electrode, and a plurality of electrodes. A step of applying a first DC signal having a first value to the sample in the flow path by using the first electrode as a working electrode and the fourth electrode as a counter electrode among the fourth electrodes different from the third electrode. Measuring a first electrical response value of the sample with respect to the first signal; and using a second electrode and a third electrode among the plurality of electrodes as a working electrode and a counter electrode, a second higher than the first value A second electric signal of the sample with respect to the second signal indicating a step of continuously applying a second DC signal having a value to the sample in the flow path for a predetermined time, and an amount of charge generated by electrolysis of moisture in the sample Measuring the response value within the predetermined time and based on the second electrical response value. Te, characterized in that it comprises a step of correcting the values obtained from the first electrical response value.

  In the second measurement method, the first electrode is used as a working electrode among the first electrode provided with a reagent containing an enzyme and the second electrode and the third electrode provided with no reagent. And applying a DC first signal having a first value to the sample in the flow path using one of the second electrode and the third electrode as a counter electrode, and the first electric power of the sample with respect to the first signal Measuring a dynamic response value, and applying a second DC signal having a second value higher than the first value to the sample in the flow path for a predetermined time using the second electrode and the third electrode as a working electrode and a counter electrode A step of continuously applying, a step of measuring a second electrical response value of the sample with respect to a second signal indicating the amount of charge generated by the electrolysis of moisture in the sample, and a second electrical Correcting a value obtained from the first electrical response value based on the response value; Characterized in that it comprises a. In the first and second measurement methods, the corrected value is treated as a measurement value of the measurement target component in the sample.

  An analysis tool applied to the first and second measurement methods is, for example, a biosensor. The sample is, for example, a biological sample, and the biological sample is a liquid sample such as blood, interstitial fluid, or urine. Components to be measured in the sample are glucose level (blood glucose level), lactate level (lactic acid level), and the like. The reagent may include an enzyme and a mediator.

  In the first and second measurement methods, the sample is preferably blood. Moreover, it is preferable that a measuring object component is glucose. The second electrical response value of the sample is preferably a value indicating hematocrit. Further, the first electrical response value of the sample is preferably a value indicating glucose before being corrected by a value indicating hematocrit.

  In the first and second measurement methods, the measurement of the first electrical response value and the measurement of the second electrical response value are performed at different timings. Either may be the order of measurement. In the first and second measurement methods, the direction of the first DC signal having the first value and the second DC signal having the second value different from the first value do not change with time, for example. Voltage, or direct current (DC) voltage. The first signal may be referred to as a first voltage, and the second signal may be referred to as a second voltage. The first value and the second value differ depending on the type of electrode material, enzyme, and mediator. However, as an absolute condition, the first value is smaller than the second value (the second value is larger than the first value).

  The applied voltage to the electrode has a lower limit of about DC 1.0 V in view of the voltage at which water electrolysis occurs. Moreover, if it is 1.0 V or more, it will become a response electric current value which is not influenced by the measuring object component in a sample. On the other hand, bubbles are generated in the sample as the applied voltage is increased, and the electrical response value is affected by the bubbles. Therefore, the upper limit of the applied voltage is about 7.0 V where no bubbles are generated. For example, when ruthenium is used as the electrode material (ruthenium electrode is used), it is preferable that the lower limit is 1.5V and the upper limit is 5.0V. When ruthenium is used as the electrode material, the DC voltage value (first value) as the first signal is, for example, about 0.5 V, and the DC voltage value (second value) as the second signal is, for example, It is about 2.5V. However, as described above, the upper limit of the second value is about 7 V where no bubbles are generated. In the present invention, the second value of the DC voltage applied to the second signal is a value of 1 V or more and 7 V or less, more preferably 1.8 or 2 V or more and 5 V or less, for example, 2.5 V. The predetermined time during which the second signal is continuously applied is preferably 2 to 5 seconds, more preferably 2.5 to 4 seconds, for example, 3 seconds. The first value of the DC voltage as the first signal is smaller than the second value, preferably 50 mV to 1.0 V, more preferably 100 mV to 750 mV, for example, 500 mV. This is a case of a ruthenium electrode. When an electrode made of a material different from ruthenium is used, an appropriate applied voltage is in a different range. Also, the appropriate applied voltage can be in a different range depending on the type of enzyme or substrate, that is, the salt.

In the first and second measurement methods, the second signal greater than or equal to a predetermined value is applied to the sample for a predetermined time or longer, and the second electrical response value is measured when the application continues for a predetermined time from the start. This is based on the following measurement principle. A second signal having a second value (for example, a DC voltage) is applied to the sample (electrode for measuring the second electrical response value) in the flow path of the analysis tool. In this case, the second electrical response value in a period of about 1 second from the start of application shows an attenuation curve depending on the concentration (for example, glucose value) of the measurement target component in the sample. On the other hand, when about 2 seconds have elapsed from the start of application, the second electrical response value does not depend on the concentration of the measurement target component, and shows a constant value corresponding to the Hct value in the sample. Such a phenomenon is caused by the following.

That is, the application of the second signal causes the charge transfer from the measurement target component to the reagent to the electrode and the charge transfer from the H ₂ O to the electrode due to the electrolysis of moisture (H ₂ O) in the sample. However, the force that moves the charge from H ₂ O to the electrode is stronger than the force that moves the charge from the component to be measured to the electrode through the reagent. When the reagent includes an enzyme and a mediator, charge transfer proceeds from the measurement target component → the reagent (enzyme → mediator) → the electrode.

For this reason, in a certain period (about 1 second) from the start of application of the second signal, the second electrical response value due to the charge transfer from the measurement target component is seen, but for a predetermined time (about 2 seconds). After the lapse of time, the charge transfer from H ₂ O to the electrode is superior to the charge transfer from the reagent to the electrode, the reagent is saturated while holding the charge, and the charge transfer from the reagent to the electrode does not occur. That is, after about 2 seconds (predetermined time) has elapsed from the start of application of the second signal, the second electrical response value to the application of the second signal is substantially the amount of charge generated by the electrolysis of H ₂ O. Is a value indicating. In other words,
The second electrical response value is a value indicating the amount of charge generated by the electrolysis of moisture in the sample, which is obtained by continuously applying the second signal for a predetermined time. When the reagent is a reagent including an enzyme and a mediator, the charge transfer from the reagent to the electrode does not occur, the charge transfer from the enzyme to the mediator does not occur, and the charge transfer from the mediator to the electrode does not occur. It means that it does not occur, and both of them.

Here, since hematocrit is the volume of formed components (blood cells) in blood, a high Hct value means that the amount of H ₂ O (plasma) to be electrolyzed is small. Therefore, Hc
The higher the t value, the lower the second electrical response value when application is continued for a predetermined time. In other words, the second electrical response value when the application continues for a predetermined time is proportional to the Hct value as a negative function. Thus, if a calibration curve (such as a table indicating a correlation) between the second electrical response value and the Hct value is created, the Hct value is calculated from the second electrical response value when the application continues for a predetermined time. can do. Such a second electrical response value or Hct value can be used to correct the measurement target component.

  From the above, the following is understood. That is, by measuring the second electrical response value when the application of the second signal has continued for a predetermined time from the start, the second electrical response value is more accurate than the second electrical response value before the predetermined time has elapsed. An electrical response value can be obtained. That is, as a result of being able to perform Hct measurement with high accuracy, it is possible to improve the accuracy of correction of the measurement target component and to measure the measurement target component with improved accuracy. Further, when the application continues for a predetermined time, the electrochemical reaction due to the application of the second signal is more stable than before the predetermined time elapses. Therefore, from the second electric response value, the second electric response before the predetermined time elapses. It is possible to obtain an Hct value with higher accuracy than the Hct value obtained from the value.

  Moreover, the analysis tool applied to the first and second measurement methods includes a plurality of electrodes. In the first measurement method, the plurality of electrodes include a first electrode provided with a reagent containing an enzyme, a second electrode and a third electrode not provided with a reagent, a first electrode, a second electrode, and A fourth electrode different from the third electrode is included. The reagent is provided on the electrode by, for example, fixing a solid reagent on the electrode.

In the measurement of the first electrical response value in the first measurement method, the first electrode is used as a working electrode and the fourth electrode is used as a counter electrode. In the measurement of the second electrical response value, the second electrode and the third electrode
An electrode is used as a working electrode and a counter electrode. Thus, an electrode without a reagent is used as a working electrode and a counter electrode used for measuring the second electrical response. The fourth electrode may or may not be provided with a reagent. However, when the reagent is provided, the accuracy of the first electrical response value is improved.

  In the second measurement method, the plurality of electrodes include at least a first electrode provided with a reagent containing an enzyme, and a second electrode and a third electrode each provided with no reagent. In the measurement of the first electrical response value in the second measurement method, the first electrode is used as a working electrode, and one of the second electrode and the third electrode is used as a counter electrode. On the other hand, in the measurement of the second electrical response value, the second electrode and the third electrode are used as a working electrode and a counter electrode. Thus, also in the second measurement method, an electrode without a reagent is used as the working electrode and the counter electrode used for measuring the second electrical response.

  In the first and second measurement methods, an electrode without a reagent is used as a working electrode and a counter electrode in the measurement of the second electrical response value. Thereby, the following advantages can be obtained. That is, in the manufacturing process of the analytical tool, for example, a liquid reagent is dropped on the electrode, or a paste-like reagent is printed and dried, thereby fixing the reagent on the electrode. At this time, the solidification state of the reagent may vary among individuals of the analytical tool, and the second electrical response value may vary among the individuals. In the first measurement method and the second measurement method, electrodes on which no reagent is provided on the working electrode and the counter electrode are used. Thereby, the dispersion | variation between individuals of a 2nd electrical response value can be suppressed.

  In the second measurement method, one of the second electrode and the third electrode is used as a working electrode in the measurement of the second electrical response value, and the other is used for the measurement of the first electrical response value and the second electrical response value. It is preferable to use it as a common counter electrode in both the measurement of the dynamic response value. However, one of the second electrode and the third electrode used as the working electrode in the measurement of the second electrical response value may be used as the counter electrode in the measurement of the first electrical response value.

  In the first and second measurement methods, two or more electrodes may be used as the working electrode and the counter electrode in the measurement of the first electrical response value and the measurement of the second electrical response value. The plurality of electrodes may include an electrode used as a reference electrode and an electrode used for measurement of measurement items other than measurement of the first electrical response value and the second electrical response value.

  Here, “the reagent is provided on the electrode” means a state where the reagent is placed in contact with the electrode, a state where the reagent is fixed, a state where the reagent is placed, or the like. In addition, although no reagent is provided on the working electrode and the counter electrode, the case where a reagent is provided between the working electrode and the counter electrode also applies to “two electrodes used for measurement of the first electrical response value”. “A reagent is provided between the working electrode and the counter electrode” means that the reagent is placed in contact with the substrate between the working electrode and the counter electrode. The case where “the reagent is provided between the working electrode and the counter electrode” includes a state where the reagent is placed in contact with the spacer or the cover that forms the flow path between the working electrode and the counter electrode. For example, the reagent is not located on the substrate, but the reagent is between the working electrode and the counter electrode when the channel is viewed in plan. In other words, “the reagent is provided between the working electrode and the counter electrode” means that the installed reagent diffuses (moves) to the measurement environment near the working electrode due to the influence of the sample introduced into the flow path. It means that it is provided as possible.

Furthermore, regarding “two electrodes used for measurement of the second electrical response value”, no reagent is provided on both of the two electrodes. In addition, regarding the two electrodes (for example, the working electrode and the counter electrode) used for measuring the second electrical response value, no reagent is provided on the working electrode and the counter electrode, but a reagent is provided between the working electrode and the counter electrode. In this case, the working electrode and the counter electrode do not apply to “two electrodes used for measuring the second electrical response value”. That is, in the measurement of the second electrical response value, it is necessary that the reagent is not provided not only in the working electrode and the counter electrode but also in the entire measurement environment of the second electrical response value.

Embodiment
Hereinafter, a liquid container according to an embodiment of the present invention will be described with reference to the drawings. The configuration of the embodiment described below is an exemplification, and the present invention is not limited to the configuration of the embodiment.

  In the following embodiments, as an example of a measurement method and a measurement apparatus, a biosensor that is an analysis tool is used to measure a glucose value in blood as a measurement target component in a sample, and an Hct value (second) in blood is measured. A measurement method and a measurement apparatus for correcting the glucose value with the electrical response value) will be described.

<Biosensor configuration>
FIG. 1A, FIG. 1B, FIG. 2A, and FIG. 2B show configuration examples of the biosensor according to the embodiment. FIG. 1A is a top view of an analysis tool (a biosensor having four electrodes) according to the embodiment, and FIG. 1B is a side view of the biosensor shown in FIG. FIG. 2A is a top view of the analysis tool (biosensor having three electrodes) according to the embodiment, and FIG. 2B is a side view of the biosensor shown in FIG.

  The biosensor 10A is applied to the first measurement method. 1A and 1B, a biosensor 10A (hereinafter “sensor 10A”) has a longitudinal direction (X direction) having one end 10a and the other end 10b, and a width direction (Y direction). Have. The sensor 10A is formed by laminating an insulating substrate 1 (hereinafter “substrate 1”), a spacer 2, and a cover 3 in the height direction (Z direction) and bonding them.

  For the substrate 1, for example, a synthetic resin (plastic) is used. Examples of synthetic resins include polyetherimide (PEI), polyethylene terephthalate (PET), polybutylene terephthalate (PBT), polyethylene (PE), polystyrene (PS), polymethacrylate (PMMA), polypropylene (PP), polyimide resin, Various resins such as acrylic resin, epoxy resin, and glass epoxy can be applied. Note that an insulating material other than synthetic resin can be applied to the substrate 1. Insulating materials include paper, glass, ceramics, biodegradable materials, etc. in addition to synthetic resins. The same material as the substrate 1 can be applied to the spacer 2 and the cover 3.

  On the upper surface of the substrate 1, as an example of a plurality of electrodes, an electrode 4, an electrode 5, an electrode 6, and an electrode 7 are provided. Each of the electrode 4, the electrode 5, the electrode 6, and the electrode 7 has a key shape having a portion extending in the width direction of the sensor 10A and a portion extending in the longitudinal direction, and the portion extending in the longitudinal direction is a lead portion 4a, The lead portion 5a, the lead portion 6a, and the lead portion 7a are formed. The lead portion 4a, the lead portion 5a, the lead portion 6a, and the lead portion 7a on the one end 10a side are not covered with the spacer 2 and the cover 3, and are used for electrical connection with the connector of the blood glucose meter 20 (FIG. 3). Is done.

  Each of the electrode 4, the electrode 5, the electrode 6, and the electrode 7 uses, for example, a metal material such as gold (Au), platinum (Pt), silver (Ag), palladium, ruthenium, or a carbon material such as carbon. Formed. For example, each of the electrode 4, the electrode 5, the electrode 6, and the electrode 7 is a metal layer having a desired thickness by forming a metal material by physical vapor deposition (PVD, for example, sputtering) or chemical vapor deposition (CVD). Can be formed as Or each of the electrode 4, the electrode 5, the electrode 6, and the electrode 7 can also be formed by printing the ink containing a carbon material on the board | substrate 1 by screen printing.

  The spacer 2 has a rectangular cutout (a recess recessed from the other end 10b toward the one end 10a) that opens toward the other end 10b. By stacking the substrate 1, the spacer 2, and the cover 3, the other end 10b of the sensor 10A is exposed by the notch surface of the spacer 2 and the notch portion (not covered by adhesion to the spacer 2). ), A space having an opening 9a formed by the upper surface of the substrate 1 provided with the electrodes and the lower surface of the cover 3 is formed. This space is used as a sample flow path 9. Air holes 11 are formed in the cover 3. The channel 9 is formed so that the sample spotted on the opening 9a is drawn (introduced) into the channel 9 by capillary action and moves toward the air hole 11 (flows in the channel 9). Has been. Part of the electrode 4, the electrode 5, the electrode 6, and the electrode 7 is exposed in the flow path 9. A reagent 8 is provided (fixed) on the electrodes 6 and 7. In contrast, the electrode 4 and the electrode 5 are not provided with a reagent.

The reagent 8 includes an enzyme and a substrate. In other words, it can be said to contain salt. The reagent 8 may further contain a mediator. The enzyme is appropriately selected according to the type of sample and the component to be measured. When the measurement target component is glucose in blood or interstitial fluid, glucose oxidase (GOD) or glucose dehydrogenase (GDH) is applied. Examples of the mediator include ferricyanide, ruthenium complex, p-benzoquinone, p-benzoquinone derivative, phenazine methosulfate, methylene blue, ferrocene, and ferrocene derivative. Among these, a ferricyanide or a ruthenium complex is preferable, and potassium ferricyanide or a ruthenium compound [Ru (NH ₃ ) ₆ ] Cl ₃ is more preferable.

  Electrode 6 and electrode 7 are used as an electrode pair used for glucose measurement. As an example, the electrode 6 is used as a working electrode, and the electrode 7 is used as a counter electrode. However, the reverse is also possible. The electrode 6 and the electrode 7 are examples of the first electrode and the fourth electrode in the first measurement method. However, when the reagent is provided on the electrode 6 and the electrode 7, the electrode 6 may be used as a counter electrode and the electrode 7 may be used as a working electrode. Further, the electrode (one of the electrode 6 and the electrode 7) used as a counter electrode in measuring the glucose value may not be provided with a reagent.

  The electrode 4 and the electrode 5 are used as an electrode pair used for measuring the Hct value. As an example, the electrode 4 is used as a working electrode, and the electrode 5 is used as a counter electrode. However, the reverse is also possible. As described above, in the examples shown in FIGS. 1A and 1B, different electrode pairs are used for the measurement of the glucose value and the measurement of the Hct value, and the electrode pair without a reagent is used for the measurement of the Hct value. used. Neither a salt component nor a mediator is provided on the electrode 4 and the electrode 5 (counter electrode for Hct value measurement).

  A biosensor 10B (hereinafter “sensor 10B”) shown in FIGS. 2A and 2B is applied to the second measurement method. The sensor 10B has a three-electrode configuration including an electrode 12, an electrode 13 and an electrode 14 instead of the electrode 4, the electrode 5, the electrode 6 and the electrode 7 in the sensor 10A. The reagent 8 is fixed on the electrode 14, the reagent is not provided on the electrode 12 and the electrode 13, and neither the salt component nor the mediator is provided.

  In measuring the glucose value, the electrode 14 is used as a working electrode, and the electrode 13 is used as a counter electrode. On the other hand, in the measurement of the Hct value, the electrode 12 is used as a working electrode and the electrode 13 is used as a counter electrode. The electrode 14 corresponds to the first electrode in the second measurement method, and the electrode 12 and the electrode 13 correspond to the second electrode and the third electrode in the second measurement method. However, the electrode 12 may be used as a counter electrode when measuring the glucose value.

<Configuration of measuring device (blood glucose level meter)>
FIG. 3A is a block diagram illustrating a configuration example of a blood glucose meter 20 that is an example of a measurement apparatus.
In FIG. 3A, the blood glucose meter 20 can be connected to, for example, a sensor 10A. The blood glucose meter 20 measures the glucose value using the connected sensor 10A and corrects the glucose value using the Hct value.

  The blood glucose meter 20 includes a first measurement unit 21, a second measurement unit 22, a switch (SW) 31, a control unit 23, a storage unit 24, and an output unit 25. The switch 31 is electrically connected to a plurality of connectors (the connector 32a, the connector 32b, the connector 32c, and the connector 32d in FIG. 3A). Connector 32a is connected to lead portion 4a (electrode 4) of sensor 10A, and connector 32b is connected to lead portion 5a (electrode 5) of sensor 10A. The connector 32c is connected to the lead portion 6a (electrode 6) of the sensor 10A, and the connector 32d is connected to the lead portion 7a (electrode 7) of the sensor 10A.

The switch 31 is a switch for switching the electrical connection between the connector 32a, the connector 32b, the connector 32c, and the connector 32d and the electrode 4, the electrode 5, the electrode 6, and the electrode 7 and the disconnection state thereof. The control unit 23 is a processor (for example, a central processing unit (CPU)) that executes a program stored in the storage unit 24. The storage unit 24 includes a memory including a main storage device such as a RAM (Random Access Memory) and an auxiliary storage device such as a ROM (Read Only Memory) and a hard disk. The storage unit 24 stores a program executed by the control unit 23, data used when the program is executed, and the like. The output unit 25 includes an output device such as a printer and a display, and communication equipment such as a signal connector and a communication interface.

  When measuring the glucose level, the control unit 23 controls the switch 31 so that the working electrode (electrode 6) and the counter electrode (electrode 7) for measuring the glucose level are electrically connected to the blood glucose meter 20. Then, the electrodes 4 and 5 are cut. The first measurement unit 21 includes a circuit, a processor, and a memory, and performs the following operation. That is, the first measurement unit 21 starts applying a DC voltage as the first signal between the electrode 6 and the electrode 7 in accordance with an instruction from the control unit 23, and after the application continues for a predetermined time, the DC voltage The response current value corresponding to the glucose value (the first electrical response value for the first signal) is measured as the response current value for. At this time, the first measurement unit 21 measures, for example, a response current value at an end point (end point) of a predetermined time during which the first signal is continuously applied as the first electrical response value. However, the time point (timing) for obtaining the first electrical response value may be a predetermined time point during the application of the first signal other than the end point of the application of the first signal.

  When measuring the Hct value, the control unit 23 controls the switch 31 so that the working electrode (electrode 4) and the counter electrode (electrode 5) for measuring the Hct value are electrically connected to the blood glucose meter 20. The electrodes 6 and 7 are cut. The second measurement unit 22 includes a circuit, a processor, and a memory, and performs the following operation. That is, the second measuring unit 22 applies a DC voltage corresponding to the second signal between the electrode 4 and the electrode 5 in accordance with an instruction from the control unit 23. The second measuring unit 22 continuously applies a DC voltage for a predetermined time from the start thereof, and sets a response current value indicating a charge amount generated by electrolysis of moisture in the sample as a second electric response. Measure as a value. However, the time point (timing) for obtaining the second electrical response value may be a predetermined time point during application of the second signal other than the end point of application of the second signal.

  The storage unit 24 stores calibration curve data for obtaining a glucose value corresponding to the first electrical response value measured by the first measurement unit 21. The control unit 23 calculates the glucose value by converting the first electrical response value into the glucose value using the calibration curve table. The glucose value is stored in the storage unit 24 or outputted (displayed or the like) from the output unit 25. Note that the calibration data for the glucose value may be a calibration curve table.

  Further, the storage unit 24 stores calibration curve data for obtaining an Hct value from the value of the second electrical response measured by the second measurement unit 22. The control unit 23 calculates the Hct value by converting the second electrical response value into the Hct value using the calibration curve data. The Hct value is stored in the storage unit 24 or output (displayed) from the output unit 25. Note that the calibration curve data for the Hct value may be a calibration curve table.

  The control unit 23 corrects the glucose value using the second electrical response value obtained from the second measurement unit 22 or the Hct value converted from the second electrical response value (hematocrit correction of the glucose value). I do. For example, the storage unit 24 stores calibration curve data and a calibration curve table that indicate the correspondence between the second electrical response value and the correction amount. The control unit 23 obtains a correction amount corresponding to the second electrical response value using the calibration curve data and the calibration curve table, performs a process of correcting the glucose value, and calculates a corrected glucose value. The corrected glucose value is stored in the storage unit 24 or output (displayed or the like) from the output unit 25. In addition, when using a 2nd electrical response value for correction | amendment of a glucose value, the structure converted into a Hct value from a 2nd electrical response value is not necessarily required. Further, the control unit 23 can perform temperature correction of the second electrical response value using calibration curve data for temperature correction and a calibration curve table.

  FIG. 3B shows a blood glucose meter 20A having a configuration corresponding to the sensor 10B having a three-electrode configuration. In this case, the connector 32d is omitted, the connector 32a is connected to the lead portion 12a (electrode 12), the connector 32b is connected to the lead portion 13a (electrode 13), and the connector 32c is connected to the lead portion 14a (electrode 14). The The switch 31 is in a state in which the first measurement unit 21, the electrode 14, and the electrode 13 are electrically connected, for example, under the control of the control unit 23 when measuring the glucose value.

  On the other hand, when measuring the Hct value, the switch 31 is in a state in which the second measurement unit 22, the electrode 12, and the electrode 13 are electrically connected, for example, under the control of the control unit 23. As described above, the switch 31 switches the connection state between the electrodes 12 to 14, the first measurement unit 21, and the second measurement unit 22.

  The 1st measurement part 21 applies DC voltage as a 1st signal between the electrode 14 as a working electrode, and the electrode 13 as a counter electrode, and measures a 1st electrical response value. The second measuring unit 22 applies a DC voltage as the second signal between the electrode 12 as the working electrode and the electrode 13 as the counter electrode, and the second electrical response value at the time when the application continues for a predetermined time from the start. Measure. Except for the above, the blood glucose meter 20 </ b> A has the same configuration as the blood glucose meter 20.

<Operation example>
FIG. 4 is a flowchart showing an operation example of the blood glucose level meter 20. In S01 shown in FIG. 4, the sample is spotted. That is, when a biosensor (for example, the sensor 10A) is connected to the blood glucose meter 20 and the opening 9a of the other end 10b of the sensor 10A is brought into contact (spotted) with blood (sample) collected from the subject, blood is caused by capillary force. Is drawn into the flow path 9 and fills the flow path 9.

  The controller 23 is applied to the electrode before spotting, observes the applied voltage, detects a change in voltage due to contact between blood and the electrode by spotting, and blood is introduced into the flow path 9. Detect that. Then, the control part 23 starts the measurement of Hct (S02).

  In S02, the control unit 23 controls the second measurement unit 22 to apply the second signal to the blood. That is, the control unit 23 instructs the second measurement unit 22 to apply a DC voltage (for example, 2.5 V) corresponding to the second signal to the electrode pair (electrode 4 and electrode 5) for Hct measurement.

The second measuring unit 22 waits for the application of the DC voltage corresponding to the second signal to continue for a predetermined time (about 2 seconds) from the start, and when the predetermined time continues, that is, the response value is (the charge amount by the reagent) The second electrical response of the blood at the time when it becomes a state showing the amount of charge generated by the electrolysis of water in the blood is measured (S03). For example, the second measurement unit 22 measures a response signal with respect to the DC voltage, performs A / D conversion, and sends the response signal to the control unit 23.

  When acquiring the second electrical response of blood to the second signal, the control unit 23 starts measuring glucose. That is, the control unit 23 applies the first signal to the blood (S04). That is, the control unit 23 instructs the first measurement unit 21 to apply a DC voltage lower than the second signal to the glucose measurement electrode pair (electrode 6 and electrode 7) as the first signal. The first measurement unit 21 applies a DC voltage according to the instruction.

  The first measurement unit 21 measures the first electrical response of blood to the first signal (S05). For example, the first measurement unit 21 measures the response current of the DC voltage applied as the first signal. The first measurement unit 21 performs A / D conversion on the response current and transmits the response current to the control unit 23.

  The control unit 23 operates as a correction unit and performs a glucose value correction process (S06). That is, the control unit 23 calculates the value of the correction target component (glucose value) contained in the blood using the value of the first electrical response (response current) acquired in S05 and the calibration curve data or the calibration curve table. To do. Further, the control unit 23 calculates the corrected glucose value using the second electrical response value (or the Hct value corresponding to the second electrical response value) acquired in S03.

  The control unit 23 outputs the corrected glucose value (S07). That is, the control unit 23 stores the glucose value corrected in S06 in the storage unit 24 and displays it on the output unit 25 (display). The control unit 23 can also transmit the glucose value to other devices via a wired or wireless network using the output unit 25. Even when the sensor 10B and the blood glucose meter 20A are applied, the same processing as in FIG. 4 is performed. For this reason, detailed description of processing is omitted. In the operation example shown in FIG. 4, the measurement of the Hct value is performed before the measurement of the glucose value.

As Example 1, the measurement of the glucose value and the measurement of the Hct value using a biosensor in which no reagent is provided on the working electrode and the counter electrode used for Hct measurement will be described. As a biosensor, a biosensor having the configuration of the sensor 10A and the electrodes 4 to 7 made of ruthenium was produced. A reagent 8 was provided on the electrodes 6 and 7. The prescription and manufacturing method of the reagent 8 are as follows. Polyvinyl alcohol (PVA146,000) 0.8 wt%, ruthenium compound _{[Ru (NH 3) 6]} Cl 3, 1.6 wt%, FAD-GDH2.7U, 1- methoxy PMS0.3 wt%, ACES buffer An enzyme solution containing (pH 6.5) was prepared. Reagent 8 was obtained by dispensing 0.5 μL of this enzyme solution and drying at 25 ° C.

  As a specimen, a mixture of whole blood of two people was used. As samples (specimen), samples having glucose values of 336 mg / dl and Hct values of 0, 20%, 42%, and 70% were prepared. Samples having an Hct value of 42% and glucose values of 0, 67 mg / dl, 600 mg / dl, and 800 mg / dl were prepared. The number n of each sample was n = 5.

The evaluation method was performed according to the following procedure.
1. Adjust the Hct value of the sample to 42%.
2. A sample added with a glucose addition solution to a sample whose Hct value was adjusted to 42% was obtained, and a sample group (sample group) having glucose values of 0 mg / dl, 67 mg / dl, 336 mg / dl, 600 mg / dl, and 800 mg / dl, respectively. 1).
3. Moreover, Hct adjustment of the sample whose glucose value was adjusted to 336 mg / dl was performed, and sample groups (referred to as sample group 2) having Hct values of 0%, 20%, 42%, and 70%, respectively, were created. 4). The Hct value and glucose value were measured under the following conditions.

(1) Measurement of second electrical response value (Hct value) 5 after connecting the electrodes (electrode 4 and electrode 5) for measuring the Hct value of the sensor 10A in which the sample is introduced into the flow path to the blood glucose meter 20 After 2 seconds (5 second open circuit), application of a DC voltage for measuring the Hct value (2.5 V: corresponding to the second signal having the second value) was started. The application of the DC voltage was continued for 2.5 seconds, and the response current (second electrical response value) after 2.5 seconds from the start of application (3 seconds from the start of measurement) was measured.

(2) Measurement of first electrical response value (glucose value) The application was stopped 2.5 seconds after the start of application of DC 2.5 V, and the circuit was opened for 7 seconds (non-energized state). During this time, switching of the electrode pair to which the DC voltage is applied (that is, switching from the electrode 4 and the electrode 5 to the electrode 6 and the electrode 7) was performed. When 7 seconds had elapsed, a DC voltage for glucose measurement (200 mV: corresponding to the first signal having the first value) was continuously applied for 5 seconds. Response current (first electrical response value) was measured at 14 seconds from the start of measurement.

Table 1 shows the measurement result of the second electrical response value of each sample having a constant glucose value (336 mg / dl) and an Hct value of 0%, 20%, 42%, and 70%. The measurement value of the second electrical response value is the measurement value (3 second value) at the time when 3 seconds have elapsed from the start of measurement shown in the measurement method of (2) above (when application has continued for 2.5 seconds). .

FIG. 5A is a graph showing the Hct linearity created based on the measurement results of Table 1, and FIG. 5B shows the second electrical response measured using the sample group 2 (constant glucose value). FIG. 5C shows the temporal change of the first electrical response value measured using the sample group 2 (constant glucose value).

  The graph of FIG. 5 (A) shows the correspondence relationship (linearity of the Hct calibration curve) between the second electrical response value and the Hct value when the glucose value is constant. As shown in FIG. 5A, the correspondence relationship between the second electrical response value and the Hct value is linear, and it was found that it can be suitably used as a calibration curve.

  From FIG. 5 (B) and FIG. 5 (C), although a response value became small, so that the Hct value was large, the substantially same waveform was observed. As shown in FIG. 5B, the second electrical response value after about 2 seconds from the start of measurement is substantially constant regardless of the difference in the Hct value (in the range of 0 to 70%). It has been found that a stable second electrical response value can be obtained even if no reagent is provided on both the working electrode and the counter electrode.

  Further, as shown in FIG. 5C, the first electrical response value shows a temporal change according to the Hct value, but is a constant value when it exceeds 13 seconds. From this, it can be seen that even if the glucose value is measured following the measurement of the Hct value, the first electrical response value that becomes a constant value after a predetermined time has elapsed can be measured.

  6A shows a temporal change in the second electrical response value measured using the sample group 1 having a constant Hct value (42%), and FIG. 6B shows a constant Hct value (42 %) Shows the time change of the first electrical response value measured using the sample group 1.

  As shown in FIG. 6 (A), regardless of the glucose value (in the range of 0 to 800 mg / dl), a temporal change in the second electrical response value having substantially the same temporal change was observed. As shown in FIG. 6 (B), the lower the glucose value, the lower the first electrical response value, but the first electrical response value that was substantially constant was observed after exceeding 13 seconds. Thus, it was found that even if the glucose value was measured following the measurement of the Hct value, a suitable glucose measurement was performed. The configurations described in the embodiments can be combined as appropriate.

DESCRIPTION OF SYMBOLS 1 ... Insulating board | substrate 2 ... Spacer 3 ... Cover 4-7, 12-14 ... Electrode 10A, 10B ... Biosensor 20, 20A ... Measuring apparatus 21 ... 1st Measuring unit 22 ... second measuring unit 23 ... control unit 24 ... storage unit 25 ... output unit 31 ... switches 32a, 32b, 32c, 32d ... connector

Claims (10)

A measurement method for measuring a measurement target component contained in a biological sample using an analysis tool in which a plurality of electrodes are arranged in a flow path,
A first electrode provided with a reagent containing an enzyme, and a second electrode and a third electrode not provided with the reagent, the first electrode, the second electrode, and the second electrode; Of the fourth electrode different from the third electrode, the first electrode is used as a working electrode and the fourth electrode is used as a counter electrode, and a DC first signal having a first value is used as the sample in the channel. Applying to:
Measuring a first electrical response value of the sample to the first signal;
A second DC signal having a second value higher than the first value is continuously applied to the sample in the flow path for a predetermined time using the second electrode and the third electrode as a working electrode and a counter electrode. And a process of
Measuring a second electrical response value of the sample to the second signal indicating the amount of charge generated by electrolysis of moisture in the sample within the predetermined time;
Correcting a value obtained from the first electrical response value based on the second electrical response value;
A measurement method comprising: A measurement method for measuring a measurement target component contained in a biological sample using an analysis tool in which a plurality of electrodes are arranged in a flow path,
Among the plurality of electrodes, among the first electrode provided with a reagent containing an enzyme and the second electrode and the third electrode not provided with the reagent, the first electrode is used as a working electrode and Applying a first DC signal having a first value to the sample in the flow path using one of the second electrode and the third electrode as a counter electrode;
Measuring a first electrical response value of the sample to the first signal;
A second DC signal having a second value higher than the first value is continuously applied to the sample in the flow path for a predetermined time using the second electrode and the third electrode as a working electrode and a counter electrode. And a process of
Measuring a second electrical response value of the sample to the second signal indicating the amount of charge generated by electrolysis of moisture in the sample within the predetermined time;
Correcting a value obtained from the first electrical response value based on the second electrical response value;
A measurement method comprising: The measurement method according to claim 1, wherein in the step of applying the first signal to the sample, the fourth electrode provided with the reagent is used as a counter electrode. The second signal is a DC voltage;
The measurement method according to claim 1, wherein the second value is a voltage value of 1 V or more and 7 V or less. The measurement method according to claim 1, wherein the sample is blood. The measurement method according to claim 1, wherein the measurement target component is glucose. The measurement method according to claim 1, wherein the second electrical response value is a value indicating hematocrit. The measurement method according to claim 7, wherein the first electrical response value is a value indicating glucose before being corrected by a value indicating the hematocrit. A first electrode provided with a reagent containing an enzyme, and a second electrode and a third electrode not provided with the reagent, respectively. In a measurement apparatus for measuring a measurement target component contained in a biological sample using an analysis tool including an electrode and a fourth electrode different from the first electrode, the second electrode, and the third electrode,
A first DC signal having a first value is applied to the sample in the flow path using the first electrode as a working electrode and the fourth electrode as a counter electrode, and the sample is applied to the sample in response to the first signal. A first measurement unit for measuring a first electrical response value;
A second DC signal having a second value higher than the first value is continuously applied to the sample in the flow path for a predetermined time using the second electrode and the third electrode as a working electrode and a counter electrode. A second measuring unit that measures a second electrical response value of the sample with respect to the second signal, the charge amount generated by electrolysis of moisture in the sample within the predetermined time;
A correction unit that corrects a value obtained from the first electrical response value based on the second electrical response value;
A measuring apparatus comprising: A first electrode including a flow path and a plurality of electrodes disposed in the flow path, wherein the plurality of electrodes are provided with a reagent containing an enzyme; a second electrode not provided with the reagent; In a measurement apparatus for measuring a measurement target component contained in a biological sample using an analysis tool including three electrodes,
The first electrode is used as a working electrode, and one of the second electrode and the third electrode is used as a counter electrode, and a DC first signal having a first value is applied to the sample in the channel. A first measurement unit for measuring a first electrical response value of the sample with respect to the first signal;
Using the second electrode and the third electrode as a working electrode and a counter electrode, a DC second signal having a second value higher than the first value is continuously applied to the sample in the channel for a predetermined time. A second measuring unit configured to measure a second electrical response value of the sample with respect to the second signal within the predetermined time period, which indicates an amount of electric charge generated by electrolysis of moisture in the sample.
A correction unit that corrects a value obtained from the first electrical response value based on the second electrical response value;
A measuring apparatus comprising:

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