Method and sensor for testing concentration of uric acid in biological sample

Abstract

The invention discloses a method and a sensor for testing the concentration of uric acid in a biological sample, and belongs to the technical field of uric acid detection. The method for testing the uric acid concentration in the biological sample comprises the following steps that an electrode set is made to make contact with the biological sample, the electrode set comprises a working electrode and a counter electrode, an excitation signal sequence is input to the working electrode, the excitation signal sequence comprises at least two continuous or discontinuous input signals, and the counter electrode is connected with the working electrode; each input signal is a constant value; at least one output signal sequence responding to the excitation signal sequence is measured, and the uric acid concentration in the biological sample is obtained according to the output signal sequence. The method is short in test time and high in HCT interference influence resistance, current signals can be effectively corrected by inputting the excitation signal sequence into the working electrode, the influence of HCT on a test result is reduced or eliminated, and then the accuracy and precision of the test result are improved.

Description

Technical Field

[0001] This application relates to a method and a sensor for testing the uric acid concentration in a biological sample, and belongs to the technical field of uric acid detection. Background Art

[0002] When detecting and analyzing an analyte in a biological sample, an input signal is applied to the working electrode of the sensor by a measuring device. The working electrode transmits the input signal into the biological sample, and the analyte in the biological sample undergoes an oxidation-reduction reaction to generate an output signal in response to the input signal, so as to determine the concentration of the analyte in the biological sample according to the output signal. The analyte includes substances such as blood glucose, blood ketone, blood lactic acid, cholesterol, uric acid, triglyceride, coagulation factor, anticoagulation factor, etc. The test results of the above analytes can be used for medical diagnosis and treatment of physiological abnormalities. For example, individuals with hyperuricemia can use a biosensor system to measure the blood uric acid level to adjust their diet and / or medication.

[0003] There are many test interferences in biosensors. For example, the hematocrit percentage (% - HCT) is a relatively obvious interference factor. This interference comes from the fact that during the diffusion of the mediator after the reaction to the electrode, its diffusion is hindered by blood cells, so that the diffusion coefficient changes due to % - HCT. And the current in the electrochemical reaction will be affected by different % - HCT. This is reflected in the factory batch calibration of the sensor test strip centered on 42% HCT. If the user's % - HCT is higher than 42%, the test will show a negative deviation, and if the user's % HCT is lower than 42%, the test will show a positive deviation. Other interference factors such as the aging effect generated during the storage time of the test strip, the test sensitivity declines relatively, or more oxidizable substances that can provide background signals are generated; there are also interferences such as the test ambient temperature and ambient humidity.

[0004] In monitoring and reducing chemical interferents, traditional biosensors basically input a single voltage on a blank electrode to measure the total possible interferent signal, so as to make a compensation for excluding interferents for the signal of the working electrode through an algorithm. However, in many cases, some possible chemicals will interfere with the working electrode to generate incorrect electrical signals, affecting the accuracy and precision of the detection. Therefore, it is necessary to develop a uric acid test method with strong anti-interference ability. Summary of the Invention

[0005] In order to solve the above problems, a method for testing the uric acid concentration in a biological sample is provided. This method has a short test time and strong anti-HCT interference effect. By inputting an excitation signal sequence to the working electrode, it can effectively correct the current signal, reduce or eliminate the influence of HCT on the test result, and further improve the accuracy and precision of the test result.

[0006] According to one aspect of the present application, a method for testing the uric acid concentration in a biological sample is provided, including the following steps:

[0007] Bring an electrode group into contact with the biological sample, wherein the electrode group includes a working electrode and a counter electrode, and input an excitation signal sequence to the working electrode, wherein the excitation signal sequence includes at least two consecutive or non-consecutive input signals, and each input signal is a constant value;

[0008] Measure at least one output signal sequence in response to the excitation signal sequence, and obtain the uric acid concentration in the biological sample according to the output signal sequence.

[0009] The excitation signal sequence can be continuous or discontinuous. In both cases, an output signal can be obtained based on the input signal, and by processing the output signal, the uric acid concentration in the biological sample can be obtained through multi-parameter compensation linear regression fitting, thereby reducing or eliminating the influence of HCT on the uric acid result and improving the accuracy of the detection result.

[0010] Optionally, the excitation signal sequence includes 4 - 6 consecutive input signals.

[0011] Optionally, the voltage value of the latter input signal in the excitation signal sequence is higher than that of the previous input signal, and the voltage range is 0.15 - 1.2V;

[0012] Preferably, the excitation signal sequence includes 5 consecutive input signals, and the voltage values of the 5 consecutive input signals are 0.15V, 0.4V, 0.6V, 0.8V, and 1.2V respectively.

[0013] Optionally, the application time of each input signal in the excitation signal sequence is not greater than 1s. The application time of each input signal can be equal or unequal, and in both cases, a corresponding output signal can be obtained and the output signal can be processed to obtain a calculation equation for the uric acid concentration.

[0014] Preferably, the application time of each input signal in the excitation signal sequence is equal.

[0015] Optionally, the number of output signals in response to each input signal is 2 - 6, preferably 5.

[0016] Optionally, the working electrode is a blank electrode, and the counter electrode is covered with an enzyme solution, and the enzyme solution includes VC reaction enzyme.

[0017] Covering the counter electrode with VC reaction enzyme can react with VC in the blood first, avoiding the interference of VC in the blood on the test result and further improving the detection accuracy; since the working voltage is applied to the working electrode, the working electrode is not covered with VC enzyme to avoid generating interference current.

[0018] Optionally, the enzyme solution further includes trehalose, PB buffer solution and CMC polymer.

[0019] According to another aspect of the present application, a method for calculating the uric acid concentration in a biological sample is provided, which makes an electrode group contact the biological sample, wherein the electrode group includes a working electrode and a counter electrode, and an excitation signal sequence is input to the working electrode, wherein the excitation signal sequence includes at least two consecutive or non-consecutive input signals, and each input signal is a constant value; the output signals in response to each input signal are 2-6;

[0020] The uric acid U 校准 The calculation method of the concentration is: U 校准 = U 计算 / (1 + f), U 计算 = a·i 11 ·i 21 / i 平均 + b, where

[0021] f is an error compensation factor, and the expression is as follows:

[0022]

[0023] where the value range of a is 300-350, the value range of b is -450 to -350, i 11 is the first output signal of the first input signal, i 21 is the first output signal of the second input signal;

[0024] The value range of K is -1 to 0, m represents the mth input signal, n represents the nth output signal in response to the mth input signal, M is any integer selected from 4-6, and N is any integer selected from 2-6; c x The value range of is -15 to 15, and the value range of x is an integer between 1 and the product of m and n; d y The value range of is -1 to 1, and the value range of y is any integer between 1 and the product of m and (n - 1); e z The value range of is -10 to 10, and the value range of z is any integer between 1 and the product, and × is the Cartesian product.

[0025] i mn is the current value in response to the input signal, It is the ratio between the endpoint currents. The error compensation factor is calculated by obtaining the current values, the ratio between the endpoint currents, and the product of the ratios between any two endpoint currents. By correcting the uric acid concentration with the error compensation factor, the uric acid concentration in the biological sample can be accurately calculated, improving the accuracy of the test results and saving the detection time.

[0026] Optionally, the voltage value of the latter input signal in the excitation signal sequence is higher than that of the previous input signal, and the voltage range is 0.15 - 1.2V.

[0027] Preferably, the excitation signal sequence includes 5 consecutive input signals, and the voltage values of the 5 consecutive input signals are 0.15V, 0.4V, 0.6V, 0.8V, and 1.2V respectively.

[0028] Optionally, the application time of each input signal in the excitation signal sequence is not more than 1s.

[0029] Preferably, the application time of each input signal in the excitation signal sequence is equal.

[0030] According to another aspect of the present application, a sensor for testing the uric acid concentration in a biological sample is provided, including:

[0031] A biosensor, the biosensor includes a working electrode, wherein the working electrode is a blank electrode or covered with a chemical agent that cannot oxidize uric acid, and the working electrode can receive an excitation signal sequence, the excitation signal sequence includes at least two consecutive or non - consecutive input signals, and each input signal is a constant value.

[0032] An electrochemical sensor, the electrochemical sensor includes a counter electrode, wherein the counter electrode is covered with a chemical agent that can oxidize uric acid.

[0033] Measure at least one output signal sequence in response to the excitation signal sequence, and obtain the uric acid concentration in the biological sample according to the output signal sequence.

[0034] The beneficial effects of the present application include but are not limited to:

[0035] 1. The method for testing and calculating the uric acid concentration in a biological sample in the present application uses a continuous or discontinuous excitation signal sequence for blood uric acid testing. The testing time is short, and the anti - HCT interference effect is strong. When calculating, a multi - parameter compensation linear regression fitting is used to calculate the uric acid concentration, which can effectively correct the current signal, reduce or eliminate the influence of HCT on the test results, and thus improve the accuracy and precision of the test results.

[0036] 2. The method of the present application for testing and calculating the uric acid concentration in a biological sample replaces the traditional long-time open-circuit incubation plus voltage oxidation method. By inputting an excitation signal sequence on the working electrode, the output signal and information obtained are increased, enabling the determination of limited chemicals and the accurate calculation of the uric acid concentration.

[0037] 3. The method of the present application for testing and calculating the uric acid concentration in a biological sample, when an excitation signal sequence is input to the working electrode, after the excitation signal sequence continuously rises to a certain extent, a signal related to the HCT of the blood sample will also be generated. Thus, through these signals, compensation for the HCT effect is made, and the uric acid concentration is calculated based on multi-parameter compensation linear regression fitting, thereby achieving the effect of reducing HCT interference and providing extremely important information for the uric acid testing process.

[0038] 4. The method of the present application for testing and calculating the uric acid concentration in a biological sample can oxidize the measurable substances existing in the diffusion protection layer on the electrode within a short time by the excitation signal sequence. By directly oxidizing uric acid in the blood, the concentration of uric acid is tested, and the sensitivity of uric acid detection can be improved even without uricase, meeting the requirements of clinical tests. Description of the Drawings

[0039] The drawings described herein are used to provide a further understanding of the present application and form a part of the present application. The schematic embodiments of the present application and their descriptions are used to explain the present application and do not constitute an improper limitation of the present application. In the drawings:

[0040] Figure 1 It is the output current based on 5 consecutive excitation signal sequences related to Embodiment 1 of the present application.

[0041] Figure 2 It is the linear relationship diagram of U 计算 related to Embodiment 1 of the present application.

[0042] Figure 3 It is the output current based on 4 non-consecutive excitation signal sequences related to Embodiment 2 of the present application.

[0043] Figure 4 It is the output current based on 6 consecutive excitation signal sequences related to Embodiment 3 of the present application.

[0044] Figure 5 It is the deviation diagram before and after calibration obtained by using the method of Embodiment 1 of the present application for uric acid detection. Detailed Description of the Embodiments

[0045] The present application will be described in detail below in conjunction with the embodiments, but the present application is not limited to these embodiments.

[0046] Embodiment 1

[0047] This embodiment relates to a method for testing and calculating the uric acid concentration in a biological sample, comprising the following steps:

[0048] Step 1: Determination of basic parameters:

[0049] S1: Take the hematocrit (HCT) parameters of five in vitro blood samples, with the unit of %, and then adjust the HCT parameters to 10%, 30%, 42%, 50%, and 70% respectively;

[0050] S2: Use an automatic biochemical analyzer to test the initial uric acid concentration U of the in vitro blood samples processed in S1 生化 , with the unit of μmol / L, and the test results are 238, 425, 613, 789, and 1052 μmol / L respectively;

[0051] S3: Apply 5 consecutive input signals to the working electrode. The voltage values of the 5 consecutive input signals are 0.15 V, 0.4 V, 0.6 V, 0.8 V, and 1.2 V respectively. The application time of each input signal is 1 s, and measure the 5 output current signals in response to each input signal. The output current signals are as Figure 1 shown. Obtain 25 output currents and 20 ratios between endpoint currents. The expression for the 20 ratios between endpoint currents is:

[0052] Figure 2 where i 11 ·i 21 / i 平均 is the abscissa, where i 11 is the first output signal of the first input signal, i 21 is the first output signal of the second input signal, U 生化 is the ordinate, establish a relationship, and fit to obtain a linear equation U 计算 = a·i 11 ·i 21 / i 平均 + b, and the values of a and b can be obtained. Figure 2 After fitting, a is 312.73 and b is -434.26.

[0053] S4: According to the linear equation, calculate the uric acid concentration in the blood sample, denoted as U 计算 . Through the calculated U 计算 and the U 生化 value obtained by testing with an automatic biochemical analyzer, calculate the remaining error f according to the following formula:

[0054] f = (U 计算 —U 生化 ) / U 生化

[0055] S5: Establish a regression equation based on the product of the remaining error calculated in step S4, the current value, the ratio between the terminal currents, and the ratio between the terminal currents. In step S3, 25 output currents and 20 ratios between the terminal currents are obtained. The ratios between the terminal currents are specifically:

[0056] R12 = i 12 / i 11 、R13 = i 13 / i 11 、R14 = i 14 / i 11 、R15 = i 15 / i 11 ;

[0057] R22 = i 22 / i 21 、R23 = i 23 / i 21 、R24 = i 24 / i 21 、R25 = i 25 / i 21 ;

[0058] R32 = i 32 / i 31 、R33 = i 33 / i 31 、R34 = i 34 / i 31 、R35 = i 35 / i 31 ;

[0059] R42 = i 42 / i 41 、R43 = i 43 / i 41 、R44 = i 44 / i 41 、R45 = i 45 / i 41 ;

[0060] R52 = i 52 / i 51 、R53 = i 53 / i 51 、R54 = i 54 / i 51 、R55 = i 55 / i 51 。

[0061] Then f = K + C1 * i 11 + C2 * i 12 + C3 * i 13+ ……+ Cx *i 55 + d1*R12 + d2*R13 + d3*R14 + …… + d y *R55 + e1*R12*R12 + e2*R12*R13 …… e z *R55*R55, where x is 25, y is 20, and z is 210.

[0062] Specific values of K, C1, C2, …… C 25 , d1, d2, d3, … d 20 , e1, e2, …… e 210 are obtained according to the above regression equation, that is, the specific expression of f is determined.

[0063] Step 2: Test and calculate the uric acid concentration in the biological sample:

[0064] Bring the electrode group into contact with the biological sample. Among them, the electrode group includes a working electrode and a counter electrode. The working electrode is a blank electrode, and the counter electrode is covered with an enzyme solution. The enzyme solution includes VC reaction enzyme, trehalose, PB buffer solution, and CMC polymer. Input 5 consecutive input signals to the working electrode. The voltage values of the 5 consecutive input signals are 0.15V, 0.4V, 0.6V, 0.8V, and 1.2V respectively. The application time of each input signal is 1s. Measure the 5 output current signals in response to each input signal. The calibrated uric acid concentration, that is, the uric acid concentration in the biological sample, is obtained according to the following formula.

[0065] U 校准 = U 计算 / (1 + f), U 计算 = 312.73·i 11 ·i 21 / i 平均 - 434.26, where

[0066] f is the error compensation factor, and the expression is as follows:

[0067]

[0068]

[0069] In this formula, K, C1, C2, …… C x , d1, d2, d3, … d y , e1, e2, …… e z have been determined through step S5.

[0070] Example 2

[0071] The difference between this embodiment and Embodiment 1 is that during step S3 and the process of testing and calculating the uric acid concentration in the biological sample, 4 discontinuous input signals are applied to the working electrode. The voltage values of the 4 discontinuous input signals are 0.2V, 0.4V, 0.7V, and 1.0V respectively. There is a 1s interval between the second and the third input signals, and the application time of each input signal is 1s. 6 output current signals in response to each input signal are measured. The output current signals are as Figure 3 shown. Similarly, the corresponding equation is obtained by using the multi-parameter fitting method in Embodiment 1, and the uric acid concentration in the calibrated biological sample is calculated through the equation.

[0072] Embodiment 3

[0073] The difference between this embodiment and Embodiment 1 is that during step S3 and the process of testing and calculating the uric acid concentration in the biological sample, 6 continuous input signals are applied to the working electrode. The voltage values of the 6 continuous input signals are 0.2V, 0.4V, 0.8V, 0.4V, 1.0V, and 1.2V respectively. The application time of each input signal is 1s. 2 output current signals in response to each input signal are measured. The output current signals are as Figure 4 shown. Similarly, the corresponding equation is obtained by using the multi-parameter fitting method in Embodiment 1, and the uric acid concentration in the calibrated biological sample is calculated through the equation.

[0074] Embodiment 4

[0075] This embodiment relates to a sensor for testing the uric acid concentration in a biological sample, including a biosensor and an electrochemical sensor. The biosensor includes a working electrode. Among them, the working electrode is a blank electrode or covered with a chemical agent that cannot oxidize uric acid. The working electrode can receive an excitation signal sequence, and the excitation signal sequence includes at least two continuous or discontinuous input signals, and each input signal is a constant value. The electrochemical sensor includes a counter electrode, and the counter electrode is covered with a chemical agent that can oxidize uric acid. At least one output signal sequence in response to the excitation signal sequence is measured, and according to the output signal sequence, the uric acid concentration in the biological sample is obtained.

[0076] When using this sensor to detect uric acid in a biological sample, by applying an excitation signal sequence to the working electrode, this sequence electrolytically oxidizes the biological sample to obtain an output signal sequence in response to the excitation signal sequence. Then, through the built-in algorithm, the deviation of the hematocrit measurement is effectively reduced, and the precision of the measurement result is significantly improved. Then, through integrating the obtained multi-parameter information, error compensation is performed on the uric acid test to obtain a more accurate test result. The built-in algorithm and the method of performing error compensation with multi-parameters can adopt the methods of Embodiments 1-3.

[0077] Comparative Example 1

[0078] This comparative example uses a hospital biochemical instrument to test uric acid. The specific method is: the uricase-catalase method.

[0079] Uric acid is hydrolyzed into allantoin, carbon dioxide and hydrogen peroxide under the action of uricase. Hydrogen peroxide reacts with sodium 3,5-dichloro-2-hydroxybenzenesulfonate and 4-aminoantipyrine under the action of peroxidase to generate a quinoneimine compound, and colorimetric determination is carried out at 520 nm to obtain the content of uric acid. The concentration of uric acid in the sample is proportional to the amount of quinoneimine pigment produced.

[0080] Comparative Example 2

[0081] This comparative example uses a household portable uric acid tester to test uric acid. The specific method is: the electrochemical method. After the uric acid in the blood sample contacts the inherent special chemical substances in the test strip, a chemical reaction occurs to generate a microcurrent, and this current is measured and converted into a uric acid value by the analyzer. The magnitude of the current is proportional to the uric acid concentration.

[0082] Test Example

[0083] The initial uric acid concentration of the processed in vitro blood samples was tested using a fully automatic biochemical analyzer for blood samples with known 10%, 30%, 42%, 50%, 70% HCT parameters. The test results were 238, 425, 613, 789, 1052 μmol / L respectively, named U 生化 。

[0084] The same blood samples were detected using the method of Example 1 to obtain the deviation diagrams before and after calibration, as Figure 5 shown. Each data point represents one test. According to Figure 5 it can be seen that the error compensation factor introduced by the calculation method of this application can correct the uric acid concentration, so as to accurately calculate the uric acid concentration in biological samples, and the deviation from the theoretical detection value of the fully automatic biochemical analyzer is small.

[0085] The above five blood samples with determined hematocrit HCT parameters were detected and the uric acid concentration was calculated using the methods of the above examples and comparative examples. The test values and relative deviations in Table 1 are the averages of 5 tests. The test results are shown in Table 1 below:

[0086] Table 1

[0087]

[0088]

[0089] According to the data in Table 1, it can be seen that the method for testing the uric acid concentration in biological samples of this application can quickly and accurately calculate the uric acid in unknown biological samples, meeting the requirements of clinical tests.

[0090] As described above, these are only embodiments of the present application. The protection scope of the present application is not limited by these specific embodiments, but is determined by the claims of the present application. For those skilled in the art, various modifications and changes can be made to the present application. Any modification, equivalent replacement, improvement, etc. made within the technical idea and principle of the present application shall be included within the protection scope of the present application.

Claims

1. A method for testing the uric acid concentration in a biological sample, characterized in that, Comprising the following steps: Bringing an electrode group into contact with a biological sample, wherein the electrode group includes a working electrode and a counter electrode, and inputting an excitation signal sequence to the working electrode, wherein the excitation signal sequence includes at least two consecutive or non-consecutive input signals, and each input signal is a constant value; Measuring at least one output signal sequence in response to the excitation signal sequence, and obtaining the uric acid concentration in the biological sample according to the output signal sequence.

2. The method according to claim 1, wherein The excitation signal sequence includes 4-6 consecutive input signals.

3. The method according to claim 1, wherein The voltage value of the subsequent input signal in the excitation signal sequence is higher than that of the previous input signal, and the voltage range is 0.15-1.2V; Preferably, the excitation signal sequence includes 5 consecutive input signals, and the voltage values of the 5 consecutive input signals are 0.15V, 0.4V, 0.6V, 0.8V, and 1.2V respectively.

4. The method according to claim 1, characterized in that, The application time of each input signal in the excitation signal sequence is not more than 1s; Preferably, the application time of each input signal in the excitation signal sequence is equal.

5. The method according to claim 1, wherein The number of output signals in response to each input signal is 2-6, preferably 5.

6. The method according to claim 1, wherein The working electrode is a blank electrode, and the counter electrode is covered with an enzyme solution, and the enzyme solution includes VC reaction enzyme.

7. A method for calculating the uric acid concentration in a biological sample, characterized in that, Bringing an electrode group into contact with a biological sample, wherein the electrode group includes a working electrode and a counter electrode, and inputting an excitation signal sequence to the working electrode, wherein the excitation signal sequence includes at least two consecutive or non-consecutive input signals, and each input signal is a constant value; the number of output signals in response to each input signal is 2-6; The uric acid U 校准 is calculated as follows: U 校准 = U 计算 / (1 + f), U 计算 = a·i 11 ·i 21 / i 平均 + b, where f is an error compensation factor, and its expression is as follows: Among them, the value range of a is 300 - 350, the value range of b is -450 to -350, i 11 is the first output signal of the first input signal, i 21 is the first output signal of the second input signal; The value range of K is from -1 to 0, m represents the m-th input signal, n represents the n-th output signal in response to the m-th input signal, M is selected from any integer in 4 - 6, and N is selected from any integer in 2 - 6; c x The value range of is from -15 to 15, and the value range of x is an integer between 1 and the product of m and n; d y The value range of is from -1 to 1, and the value range of y is any integer between 1 and the product of m and (n - 1); e z The value range of is from -10 to 10, and the value range of z is from 1 to any integer between the products of, and × is the Cartesian product.

8. The method according to claim 7, wherein The voltage value of the subsequent input signal in the excitation signal sequence is higher than that of the previous input signal, and the voltage range is 0.15-1.2V; Preferably, the excitation signal sequence includes 5 consecutive input signals, and the voltage values of the 5 consecutive input signals are 0.15V, 0.4V, 0.6V, 0.8V, and 1.2V respectively.

9. The method according to claim 7, characterized in that, The application time of each input signal in the excitation signal sequence is not more than 1s; Preferably, the application time of each input signal in the excitation signal sequence is equal.

10. A sensor for testing the uric acid concentration in a biological sample, characterized in that, Including: A biosensor, the biosensor includes a working electrode, wherein the working electrode is a blank electrode or covered with a chemical agent that cannot oxidize uric acid, and the working electrode can receive an excitation signal sequence, and the excitation signal sequence includes at least two consecutive or non-consecutive input signals, and each input signal is a constant value; An electrochemical sensor, the electrochemical sensor includes a counter electrode, wherein the counter electrode is covered with a chemical agent that can oxidize uric acid; Measuring at least one output signal sequence in response to the excitation signal sequence, and obtaining the uric acid concentration in the biological sample according to the output signal sequence.

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