Analytical instrument for measuring creatinine

Abstract

The invention provides an analytical instrument for detecting creatinine content. The analytical instrument comprises a positive voltage generation circuit connected with a working electrode lead and a voltage switching circuit connected with a counter electrode lead, the measurement process comprises the following steps: adding a sample into the biosensor, applying positive voltage to the working electrode and the counter electrode, then applying negative voltage, and calculating the content of an analyte in the sample according to obtained electrical data. According to the invention, the resolution of the measured current signal can be obviously improved, so that the detection precision and accuracy of measurement are improved.

Description

Technical Field

[0001] The invention belongs to the field of electrochemical detection, and in particular relates to a measuring method and an analytical instrument for measuring. Background Art

[0002] The basic structure of the biosensor includes an insulating base plate, an electrode system located on the insulating base plate, a reaction reagent disposed on the electrode system, and the electrode system includes a working electrode and a counter electrode, which are respectively connected to the pins of the analytical instrument through electrode leads, and a sample channel is disposed on the electrode, and the sample to be tested (such as physiological fluid) contacts the reaction reagent through the sample channel, and the analyte to be tested in the sample reacts with the reaction reagent to generate an electrical signal. The analytical instrument gives the test result based on the generated electrical signal.

[0003] The electrochemical method is a common method for biosensors to measure the concentration of analytes in physiological fluids (such as blood or plasma byproducts). The analyte components are allowed to react with specific reagents and produce oxidizable (or reduced) substances proportional to the concentration of the analyte, and finally the concentration of the analyte is converted into an electrical signal that is easy to measure by the instrument, such as a current signal or an impedance signal, and the concentration of the analyte is in a corresponding relationship with the signal size. For example, in the electrochemical measurement of creatinine, creatinine is converted into a current signal value related to the creatinine concentration under the action of reaction enzymes such as creatinase, creatinase, sarcosine oxidase, and peroxidase.

[0004] Improving the measurement resolution of the analyte to be measured is an important factor that needs to be considered in the design of biosensors and analytical instrument measurement systems. For example, the resolution of the current signal will affect the accuracy of the detection, so how to improve the resolution of the current signal of the biosensor is an important research topic in the field of biosensors.

[0005] One of the methods to improve the resolution of the electrical signal of a biosensor is to adjust the reagent formula in the biosensor. Once the reagent formula of the biosensor is determined, if you want to further change the magnitude of the current to improve the resolution of the electrical signal, the usual approach is to change the sensor's electrode structure or add a sample channel to increase the reaction area between the sample and the reagent. The above-mentioned methods all require redesigning the biosensor, which will inevitably increase the design cost and extend the product development cycle. Summary of the invention

[0006] In order to solve the above problems, the present invention provides a new electrochemical biosensor, which can improve the resolution of the current signal measured by the electrochemical method, thereby improving the detection accuracy of the electrochemical biosensor.

[0007] The present invention provides an analytical instrument for detecting creatinine content, comprising a pin connected to an electrode lead of a biosensor, the pin connected to a current-voltage conversion circuit, the current-voltage conversion circuit connected to an analog-to-digital conversion circuit, the analog-to-digital converter connected to a microcontroller, and also comprising a positive voltage generating circuit connected to a working electrode lead, and a voltage switching circuit connected to a counter electrode lead; during measurement, a voltage is applied to a working electrode and a counter electrode of the biosensor, so that the voltage difference between the working electrode and the counter electrode is a positive voltage, and the duration is T1 seconds, or no voltage is applied and the duration is T1 seconds; after T1 seconds, a voltage is applied between the working electrode and the counter electrode of the biosensor, so that the voltage difference between the working electrode and the counter electrode is a negative voltage, and the duration is T2 seconds; and the content of an analyte in a sample is calculated based on electrical data during T2 seconds.

[0008] Furthermore, a voltage is applied to the working electrode and the counter electrode of the biosensor for a period of T1 seconds, so that the voltage of the working electrode is 0.4 V higher than the voltage of the counter electrode.

[0009] More specifically, a voltage of 0.4 V was applied to the working electrode, and a voltage of 0 V was applied to the counter electrode.

[0010] Furthermore, a voltage is applied to the working electrode and the counter electrode of the biosensor for a period of T2 seconds, so that the voltage of the working electrode is 0.4 V lower than the voltage of the counter electrode.

[0011] More specifically, a voltage of 0.4 V was applied to the working electrode, and a voltage of 0.8 V was applied to the counter electrode.

[0012] Furthermore, in the calculation step, the content of the analyte in the sample is calculated based on the measured value at time T2.

[0013] Furthermore, the T1 time is greater than or equal to 30S, and the T2 time is greater than or equal to 3S.

[0014] Furthermore, the reagents on the biosensor include creatinase, creatinase, sarcosine oxidase, peroxidase and a reduced electron mediator.

[0015] The present invention also provides a new measurement method applied to an electrochemical biosensor, by which the resolution of the current signal measured by the electrochemical method can be improved, thereby improving the detection accuracy of the electrochemical biosensor.

[0016] Specifically, the present invention provides a measurement method, which uses an electrochemical biosensor for detection. The electrochemical biosensor includes a working electrode and a counter electrode. The measurement process includes:

[0017] Step 1: Add sample to the biosensor;

[0018] Step 2: signal incubation, duration T1 seconds;

[0019] Step 3: After T1 seconds have passed, a voltage is applied between the working electrode and the counter electrode of the biosensor for a duration of T2 seconds;

[0020] Step 4: Based on the electrical data obtained in step 3, calculate the content of the analyte in the sample.

[0021] Furthermore, in step 2, a voltage is applied between the working electrode and the counter electrode of the biosensor so that the voltage difference between the working electrode and the counter electrode is a positive voltage, zero or a negative voltage.

[0022] Furthermore, in step 3, a voltage is applied between the working electrode and the counter electrode of the biosensor to generate a voltage difference between the working electrode and the counter electrode. The voltage difference generated in step 3 is opposite to the voltage difference generated in step 2.

[0023] The voltage difference generated in step 3 is opposite to the voltage difference generated in step 2, which means that when the voltage difference generated between the working electrode and the counter electrode in step 2 is a positive voltage, the voltage difference generated between the working electrode and the counter electrode in step 3 is a negative voltage.

[0024] Furthermore, in step 4, the content of the analyte in the sample is calculated based on the measurement value at time T2.

[0025] Furthermore, step 1 also includes determining whether the amount of sample added is sufficient. After adding the sample to the biosensor, a voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage difference between the working electrode and the counter electrode is a positive voltage. When a current change is detected, it is determined that the amount of sample added meets the requirements and is sufficient, and then step 2 is entered.

[0026] More specifically, the present invention provides a method for measuring creatinine, which is detected by an electrochemical biosensor, wherein the electrochemical biosensor includes a working electrode and a counter electrode, and the measurement process includes:

[0027] Step 1: Add sample to the biosensor;

[0028] Step 2: applying voltage to the working electrode and the counter electrode of the biosensor so that the voltage difference between the working electrode and the counter electrode is a positive voltage, or not applying voltage; duration T1 second;

[0029] Step 3: After T1 seconds have passed, a voltage is applied between the working electrode and the counter electrode of the biosensor so that the voltage difference between the working electrode and the counter electrode is a negative voltage, which lasts for T2 seconds;

[0030] Step 4: Based on the electrical data obtained in step 3, calculate the content of the analyte in the sample.

[0031] Furthermore, in step 2 of the method for measuring creatinine, a voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage of the working electrode is 0.4 V higher than the voltage of the counter electrode.

[0032] More specifically, in step 2 of the method for measuring creatinine, a voltage of 0.4 V is applied to the working electrode, and a voltage of 0 V is applied to the counter electrode.

[0033] Furthermore, in step 3 of the method for measuring creatinine, voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage of the working electrode is 0.4 V lower than the voltage of the counter electrode.

[0034] More specifically, in step 3 of the method for measuring creatinine, a voltage of 0.4 V was applied to the working electrode, and a voltage of 0.8 V was applied to the counter electrode.

[0035] Furthermore, in step 4 of the method for measuring creatinine, the content of the analyte in the sample is calculated based on the measurement value at time T2.

[0036] Furthermore, the creatinine measurement method also includes a step of determining whether to inject a sample. Specifically, in step 1 of the creatinine measurement method, a voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage difference between the working electrode and the counter electrode is a positive voltage. Specifically, when a current change is detected, it indicates that the sample has entered the reaction area of ​​the biosensor. Subsequently, step 2 is entered, and the analysis instrument starts T1 timing.

[0037] Furthermore, in the sample injection judgment step of the creatinine measurement method, a voltage of 0.4V is applied to the working electrode, and a voltage of 0V is applied to the counter electrode.

[0038] Furthermore, in the method for measuring creatinine, the T1 time is greater than or equal to 30 seconds, and the T2 time is greater than or equal to 3 seconds.

[0039] Furthermore, the biosensor for creatinine detection includes detection reagents, wherein the reagents include creatinase, creatinase, sarcosine oxidase, peroxidase and a reduced electron mediator.

[0040] The present invention adds a signal incubation step between the conventional sample addition step and the signal measurement and calculation step, so that the present invention enters the conventional signal measurement and result calculation step only after waiting for a certain time (T1s) after the sample addition step.

[0041] The signal incubation described in the present invention can be carried out in a manner where no voltage is applied between the working electrode and the counter electrode, or after the voltage is applied, the voltage difference between the working electrode and the counter electrode is 0; it can also be carried out in a manner where a voltage is applied to the working electrode and the counter electrode, and the voltage difference generated between the working electrode and the counter electrode after the voltage is applied is opposite to the voltage difference in the signal measurement stage.

[0042] The beneficial effects of the present invention are:

[0043] The signal incubation step is added to the measurement method of the present invention. By waiting for a certain period of time and increasing the voltage, the current signal resolution of the electrochemical method measurement is significantly improved without changing the biosensor test paper formula and increasing the sample contact chamber (sample channel), thereby improving the measurement accuracy. The method of the present invention is applied to the electrochemical measurement of creatinine, blood sugar, uric acid, cholesterol, triglycerides and other intermediate reactions that produce H2O2, and high-resolution electrical signals and high detection accuracy can be achieved. At the same time, the research and development cost of the biosensor can also be effectively reduced. BRIEF DESCRIPTION OF THE DRAWINGS

[0044] Figure 1 It is a schematic diagram of the test circuit of the present invention.

[0045] Figure 2 It is a waveform diagram of the voltage applied across the working electrode and the counter electrode.

[0046] Figure 3 It is a schematic diagram of a biosensor for creatinine detection in which the electrode is covered with an insulating layer but not with a gap layer and a covering layer.

[0047] Figure 4 is a schematic diagram of the exploded view of the biosensor used for creatinine detection.

[0048] Figure 5 is a cross-sectional view of a biosensor for creatinine detection.

[0049] Figure 6 It is a measurement flow chart of the present invention.

[0050] Figure 7 This is a graph showing the relationship between creatinine concentration and current signal size at different incubation voltages and times.

[0051] Figure 8 It is the relationship between creatinine concentration and current signal size when the incubation voltage is the same and the incubation time is different.

[0052] Fig. 9 It is the waveform diagram of the current signal changing with time during the signal testing phase. DETAILED DESCRIPTION

[0053] The present invention is described in detail below with reference to specific embodiments.

[0054] Example 1 Biosensor and Analytical Instrument for Detecting Creatinine Content

[0055] like Figure 3 , Figure 4 and Figure 5 The biosensor 100 for detecting creatinine content in a blood sample shown in the figure includes an insulating substrate 103, a working electrode 101 and a counter electrode 102 are arranged on the insulating substrate 103, and an electrode lead 104 connected to the working electrode or the counter electrode, and the other end of the electrode lead is connected to an analytical instrument. An insulating layer 105 is used as a reaction area forming layer to cover the electrode, and an opening is formed at the position of the signal reaction area 110 of the electrode corresponding to the insulating layer, and a reaction reagent is added into the opening. A sample channel 108 is formed between the insulating layer 105 and the covering layer 106 through a gap layer 107, and a pore 109 is arranged on the covering layer. Among them, Figure 3 The schematic diagram of the biosensor electrode is covered with an insulating layer, but not with a gap layer and a covering layer, which shows the arrangement of the electrodes under the insulating layer. The signal reaction area 110 contains a reaction reagent, and the reagent components include creatinase, creatinase, sarcosine oxidase, peroxidase, enzyme mediator, surfactant and buffer.

[0056] The reaction principle of the biosensor for detecting creatinine is as follows: insert the biosensor 100 for detecting the creatinine content of the blood sample into the analytical instrument. When the blood sample enters the biosensor through the sample channel 108, the creatinine in the blood sample generates creatine under the action of creatinase and creatinase in the biosensor (Formula (1)); creatine, H2O and O2 react under the action of creatine oxidase to generate formaldehyde, glycine and H2O2 (Formula (2)); the reduced electron mediator and H2O2 react under the action of peroxidase to generate oxidized electron mediator and H2O (Formula (3)); after the measuring circuit applies a potential, the oxidized electron mediator absorbs electrons to react and generates a reduced electron mediator (Formula (4)). In Formula (4), the number of electrons is positively correlated with the creatinine concentration. The creatinine measuring and analyzing instrument can convert the detected current signal into a specific value of the creatinine concentration through a solidified software system.

[0057]

[0058]

[0059]

[0060]

[0061] The test circuit of the creatinine measurement and analysis instrument is as follows Figure 1As shown, it includes a metal shrapnel 1 connected to the working electrode 101 of the biosensor 100 and a metal shrapnel 2 connected to the counter electrode 102 of the biosensor. The metal shrapnel 1 and the metal shrapnel 2 are respectively connected to the current-voltage conversion circuit 3, and the current-voltage conversion circuit 3 is composed of a transimpedance amplifier. The metal shrapnel can also be called a pin. The current-voltage conversion circuit 3 is connected to the analog-to-digital conversion circuit 5, and the analog-to-digital converter 5 is connected to the microcontroller 7. The microcontroller 7 is composed of a single-chip microcomputer as a core, which is used to control the 0.8V or 0V voltage switching circuit 6 and collect the test signal of the analog-to-digital conversion circuit 5 and calculate the test result. The microcontroller is provided with a data storage unit 8. The test circuit also includes a 0.4V voltage generating circuit 4 and a 0.8V or 0V voltage switching circuit 6. The 0.4V voltage generating circuit 4 is generated by dividing the voltage of the reference source and following the output through the operational amplifier. The 0.8V voltage in the 0.8V or 0V voltage switching circuit 6 is generated by dividing the voltage of the reference source and following the output through the operational amplifier, and 0V is directly connected to the system ground, and the switching between the two is realized by switching the analog switch.

[0062] The voltage value of the voltage generating circuit 4 and the voltage value of the voltage switching circuit 6 can be set according to actual conditions, and are not necessarily 0.4V or 0.8V.

[0063] Example 2 Method for measuring creatinine content in blood samples

[0064] The method for measuring the creatinine content in a blood sample comprises the following steps.

[0065] Step 1: Sample injection detection: Insert the biosensor into the analytical instrument and add the sample to the biosensor. The analytical instrument detects whether the sample has entered the reaction area of ​​the biosensor to determine whether to inject the sample. When the instrument detects the injection, the instrument enters the signal incubation period.

[0066] Step 2: Signal incubation period: During the incubation period, voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage difference between the working electrode and the counter electrode is a positive voltage (i.e., the working electrode voltage is higher than the counter electrode voltage), which lasts for T1 seconds, and then enters the signal measurement period.

[0067] Step 3: Signal measurement period: During the measurement period, a voltage is applied between the working electrode and the counter electrode of the biosensor so that the voltage difference between the working electrode and the counter electrode is a negative voltage (i.e., the working electrode voltage is lower than the counter electrode voltage). At this time, a current signal is generated between the working electrode and the counter electrode. The current signal is converted into a voltage signal through a current-voltage conversion circuit, and the signal is sampled and recorded through an analog-to-digital conversion circuit. The duration is T2 seconds.

[0068] Step 4: Calculation of results: Take the measured value at time T2 seconds during the measurement period and convert it into creatinine concentration using the calibration curve.

[0069] The specific durations of the durations T1 seconds and T2 seconds can be determined as needed based on experimental results, for example, they are ideal times selected after repeated laboratory tests of the biosensor.

[0070] like Figures 1 to 6 As shown, the biosensor 100 of Example 1 and the test circuit of the analytical instrument are used to measure the creatinine content in the blood sample.

[0071] During the sampling detection stage, a blood sample is added to the sampling channel 108 of the biosensor. The test circuit of the analytical instrument applies a voltage of 0.4V to the working electrode 101 and a voltage of 0V to the counter electrode 102. At this time, the analog-to-digital conversion circuit 5 is always working. When the controller 7 detects a current change through the analog-to-digital conversion circuit 5, it indicates that the sample has entered the reaction area 110. At this time, the controller starts the T1 timing and the measurement enters the signal incubation stage.

[0072] In the signal incubation stage, when the addition of a sample is detected, the incubation program is started. In this embodiment, a positive voltage incubation method is adopted, that is, after the voltage is applied, the voltage difference between the working electrode and the counter electrode is a positive voltage. Specifically, the voltage applied to the working electrode 101 is 0.4V, and the voltage applied to the counter electrode 102 is 0V. At this time, the working electrode voltage is 0.4V relative to the counter electrode voltage. The incubation voltage waveform is as follows: Figure 2 As shown in the T1 time period, T1 represents the application of a 0.4V incubation voltage between the working electrode and the counter electrode, which lasts for T1 seconds. In this process, the analog-to-digital conversion circuit 5 does not need to work. When the controller 7 reaches the time of T1 seconds, the incubation ends and the signal measurement phase is entered. In this embodiment, the current signal generated during the incubation phase does not participate in the calculation of the result. In another embodiment, the analog-to-digital conversion circuit 5 works, but the current signal generated during the incubation phase does not participate in the calculation of the result.

[0073] In the signal measurement stage, when the signal incubation is completed, the signal measurement program is started. In this embodiment, a negative voltage is used for measurement, that is, after the voltage is applied, the voltage difference between the working electrode and the counter electrode is a negative voltage. Specifically, a voltage of 0.4V is applied to the working electrode, and the voltage applied to the counter electrode is controlled by the controller and switched from 0V to 0.8V. At this time, the working electrode voltage is -0.4V relative to the counter electrode voltage, and the waveform is as follows: Figure 2 As shown in the T2 time period, T2 represents the -0.4V measurement voltage applied between the working electrode and the counter electrode, and the duration is T2 seconds. The controller 7 continuously collects the voltage signal generated by the creatinine current signal flowing through the current-voltage conversion circuit 3 through the analog-to-digital conversion circuit 5, and stores it in the storage unit 8 of the controller 7. When the T2 second time is reached, the measurement is stopped. Enter the result calculation stage.

[0074] In the result calculation stage, the controller 7 analyzes the data collected within T2 seconds, obtains the measured value at the time T2 seconds of the measurement period, and converts it into creatinine concentration through the calibration curve.

[0075] Whether a voltage difference is a positive voltage or a voltage difference is a negative voltage is relative.

[0076] Taking the voltage of the counter electrode as a reference, a voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage of the working electrode is higher than that of the counter electrode, and the voltage difference between the two is a positive voltage. For example, when the voltage applied to the working electrode is 0.4V and the voltage applied to the counter electrode is 0V, the voltage difference between the working electrode and the counter electrode is 0.4V, that is, the voltage of the working electrode relative to the counter electrode is 0.4V. When a voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage of the working electrode is lower than that of the counter electrode, the voltage difference between the two is a negative voltage. For example, when the voltage applied to the working electrode is 0.4V and the voltage applied to the counter electrode is 0.8V, the voltage difference between the working electrode and the counter electrode is -0.4V, that is, the voltage of the working electrode relative to the counter electrode is -0.4V.

[0077] On the contrary, with the voltage of the working electrode as the reference, a voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage of the working electrode is higher than that of the counter electrode, and the voltage difference between the two is a negative voltage. For example, when the voltage applied to the working electrode is 0.4V and the voltage applied to the counter electrode is 0V, the voltage difference between the working electrode and the counter electrode is -0.4V, that is, the voltage of the working electrode relative to the counter electrode is -0.4V. When a voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage of the working electrode is lower than that of the counter electrode, the voltage difference between the two is a positive voltage. For example, when the voltage applied to the working electrode is 0.4V and the voltage applied to the counter electrode is 0.8V, the voltage difference between the working electrode and the counter electrode is 0.4V, that is, the voltage of the working electrode relative to the counter electrode is 0.4V.

[0078] Example 3 Relationship between creatinine concentration and current signal size under different incubation voltages and times

[0079] In order to compare the relationship between the creatinine concentration in the sample and the current signal under different incubation voltages and incubation times, this embodiment adopts three groups of experimental schemes to carry out comparative experiments.

[0080] Experimental Group 1

[0081] In this experimental group, the measurement steps include step 1 sample injection detection, step 3 signal measurement and step 4 result calculation in Example 2. This experimental group does not include the signal incubation in Example 2. Specifically, after completing the sample injection detection according to the method of Example 2, directly enter the signal measurement stage, apply voltage to the working electrode and the counter electrode of the biosensor, even if the working electrode voltage is -0.4V relative to the counter electrode voltage, the duration is T2 seconds, enter the result calculation stage, and take the measurement value at T2 seconds to calculate the creatinine concentration.

[0082] Experimental Group 2

[0083] In this experimental group, the measurement steps include step 1 sampling detection, step 2 signal incubation, step 3 signal measurement and step 4 result calculation in Example 2, except that the signal incubation is a non-pressure incubation. Specifically, after completing the sampling detection according to the method of Example 2, the signal incubation stage is entered, and a non-pressure incubation method is adopted in this stage, for example, the voltage difference between the working electrode and the counter electrode is 0V. When the controller timing T1 reaches 30s (i.e., incubation without pressure for 30s), the signal measurement is entered, and a constant -0.4V voltage is applied to the working electrode and the counter electrode of the biosensor for a duration of T2 seconds, and the result calculation stage is entered, and the measurement value at T2 seconds is taken to calculate the creatinine concentration.

[0084] Experimental Group 3

[0085] In this experimental group, the measurement steps include step 1 of sample injection detection, step 2 of signal incubation, step 3 of signal measurement and step 4 of result calculation in Example 2. Specifically, after the sample injection detection is completed according to the method of Example 2, the signal incubation stage is entered, and a voltage is applied to the working electrode and the counter electrode of the biosensor so that the working electrode voltage is 0.4V relative to the counter electrode voltage. When the controller timing T1 reaches 30s, the signal measurement is entered. In the signal measurement stage, a constant voltage of -0.4V is applied to the working electrode and the counter electrode of the biosensor for a duration of T2 seconds, and the result calculation stage is entered, and the measured value at the time of T2 seconds is taken to calculate the creatinine concentration.

[0086] The T2 time of the above three experimental groups was 3s.

[0087] The signal measurement stage of this embodiment adopts a method of applying 0.4V to the working electrode and 0.8V to the counter electrode. Of course, in other embodiments, it is also possible to apply 0.5V to the working electrode and 0.9V to the counter electrode, as long as the voltage difference between the working electrode and the counter electrode is -0.4V. Similarly, the voltage applied in the incubation stage can also be selected according to the actual situation of product design, for example, 0.5V is applied to the working electrode and 0.1V is applied to the counter electrode in the incubation stage.

[0088] In this embodiment, fresh blood is used to prepare multiple groups of samples with known creatinine concentrations. The specific concentrations of creatinine in the samples are shown in Table 1. The test results are shown in Tables 1 and Figure 7 shown. Figure 7 The figure is a relationship diagram between creatinine concentration and current signal size under different incubation voltages and times, where 0V0S represents no incubation, 0V30S represents incubation for 30 seconds but no pressure is applied during the incubation stage, and 0.4V30S represents that a 0.4V incubation voltage is applied during the incubation stage.

[0089] Experimental group 1 ( Figure 7 The test results show that the measurement slope of experimental group 1 is only 0.0002uA / uM, and there is basically no signal gradient.

[0090] Experimental group 2 ( Figure 7 The test results show that the slope of experimental group 2 is 0.0013uA / uM, which is more than 6 times the slope of experimental group 1 measured directly without incubation, and the detection signal gradient is greatly increased.

[0091] Experimental group 3 ( Figure 7 The test results show that the measured slope of experimental group 3 is 0.0019uA / uM, which is more than 40% higher than that of experimental group 2, and the detection signal gradient is further increased.

[0092] According to the above analysis, adding an incubation step during creatinine measurement can increase the detection signal gradient, especially after adding a pressure incubation step, the detection signal gradient can be greatly increased. This may be because the incubation process helps the accumulation of electrons during the reaction, especially the pressure incubation (for example, applying a 0.4V voltage for 30s), which can greatly increase the detection reaction current gradient.

[0093] Table 1

[0094]

[0095]

[0096] Example 4: Analysis of the relationship between creatinine concentration and reaction current using different incubation times

[0097] The method of this embodiment is basically the same as that of Experimental Group 3 of Example 3, except that the incubation time T1 is 10s, 30s, and 40s respectively. The test results are shown in Tables 2 and Figure 8 shown. Figure 8 The relationship between creatinine concentration and current signal at different incubation times, where 0.4V10S represents the application of 0.4V incubation voltage for 10s, 0.4V30S represents the application of 0.4V incubation voltage for 30s, and 0.4V40S represents the application of 0.4V incubation voltage for 40s. The current signal slopes corresponding to incubation times of 10S, 30S, and 40S are 0.002uA / uM, 0.0026uA / uM, and 0.0029uA / uM, respectively, and the current signal increases with the increase of incubation time. Considering the test efficiency and signal detection gradient, T1 can be selected as 30 seconds.

[0098] Table 2

[0099]

[0100] Example 5 analyzes the waveform of the current signal changing with time during the measurement phase.

[0101] This embodiment adopts the method of Experimental Group 3 of Embodiment 3, the incubation voltage is 0.4V, the incubation time T1 is 30s, the creatinine concentration of the sample under test is 227uM, and within the 30s incubation period, the system does not measure the current value of the creatinine signal. After the incubation ends, the signal measurement period begins. At this time, the system monitors the creatinine current signal. The waveform of the electrical signal changing with time is shown in FIG. Fig. 9 As shown, Fig. 9 It is a waveform diagram of the current signal value changing with time during the signal measurement period. Specifically, when the test time is 0s to 2s, it is a transition state. In this state, the current signal changes violently and is more random, so the test result is not suitable for selection in this time period. When the signal measurement period is after 2s, the current signal is in a steady state, the signal changes slowly over time, and the signal acquisition is stable, so the test result should be selected in this state. In the present embodiment, T2 selects the current signal test value at 3 seconds as the test value of the creatinine signal.

[0102] Example 6 Stability Test

[0103] This example adopts the method of Experimental Group 3 of Example 3 to test samples with different creatinine contents in Table 3. Each sample is tested 5 times. The test results are shown in Table 3.

[0104] Table 3: Five measurement results with 0.4V 30S incubation voltage

[0105]

[0106] From the data in Table 3, it can be seen that the method of the present invention, that is, after the detection sample is injected, a positive voltage of 0.4V is applied, incubated for 30s, and -0.4V is applied for signal testing to obtain the current value at time T2, and the CV value is less than 5% at 4 concentrations. Therefore, the measurement method of the present invention can meet the measurement requirements and can be used for actual measurement.

[0107] Example 7 Accuracy Test

[0108] The creatinine analysis instrument was made by the method of the present invention, and the test samples of different concentrations were tested. Each concentration sample was tested 5 times. The test results are shown in Table 4:

[0109] Table 4:

[0110]

[0111]

[0112] It can be seen from the data in Table 4 that the creatinine analyzer made by the method of the present invention has high measurement precision and good accuracy.

Claims

1. An analytical instrument for detecting creatinine content, comprising a pin connected to an electrode lead of a biosensor, the pin connected to a current-voltage conversion circuit, the current-voltage conversion circuit connected to an analog-to-digital conversion circuit, the analog-to-digital converter connected to a microcontroller, characterized in that: It also includes a positive voltage generating circuit connected to the working electrode lead and a voltage switching circuit connected to the counter electrode lead; during measurement, a voltage is applied to the working electrode and the counter electrode of the biosensor so that the voltage difference between the working electrode and the counter electrode is a positive voltage, which lasts for T1 second, or no voltage is applied and lasts for T1 second; after T1 second, a voltage is applied between the working electrode and the counter electrode of the biosensor so that the voltage difference between the working electrode and the counter electrode is a negative voltage, which lasts for T2 seconds; based on the electrical data during T2 seconds, the content of the analyte in the sample is calculated.

2. The analytical instrument according to claim 1, characterized in that During a duration of T1 seconds, a voltage is applied to the working electrode and the counter electrode of the biosensor, so that the voltage of the working electrode is 0.4 V higher than the voltage of the counter electrode.

3. The analytical instrument according to claim 2, characterized in that A voltage of 0.4 V was applied to the working electrode, and a voltage of 0 V was applied to the counter electrode.

4. The analytical instrument according to claim 1, characterized in that During a duration of T2 seconds, a voltage is applied to the working electrode and the counter electrode of the biosensor, so that the voltage of the working electrode is 0.4 V lower than that of the counter electrode.

5. The analytical instrument according to claim 4, characterized in that A voltage of 0.4 V was applied to the working electrode, and a voltage of 0.8 V was applied to the counter electrode.

6. The analytical instrument according to claim 1, characterized in that In the calculation step, the content of the analyte in the sample is calculated based on the measured value at time T2.

7. The analytical instrument according to claim 1, characterized in that The T1 time is greater than or equal to 30S, and the T2 time is greater than or equal to 3S.

8. The analytical instrument according to claim 1, characterized in that The reagents on the biosensor include creatinase, creatinase, sarcosine oxidase, peroxidase and a reduced electron mediator.

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