Method and apparatus for measuring stable hemoglobin A1c

Abstract

To provide a measurement method capable of measuring a stable hemoglobin A1c value that is closer to a true value in a specimen in which the stable hemoglobin A1c value becomes higher.SOLUTION: A measurement method of stable hemoglobin A1c by means of a separation analysis method of hemoglobin with cation exchange as a principle comprises the steps of: obtaining an analysis signal from a blood specimen being a measurement object by means of the separation analysis method; deciding a C value being a peak value of a stable hemoglobin A1c peak and an X value being a peak value of a specific peak appearing between the stable hemoglobin A1c peak and a hemoglobin A0 peak from the analysis signal; and correcting the C value by obtaining a C' value by applying the C value and the X value to a predetermined operational expression and lowering the C value. The operational expression is determined on the basis of the correlation between the C value and the X value whose stable hemoglobin A1c value is obtained from a known blood specimen by means of the separation analysis method.SELECTED DRAWING: Figure 10

Description

[Technical Field]

[0001] The present invention relates to a method and an apparatus for measuring stable hemoglobin A1c in a blood sample. [Background Art]

[0002] Regarding hemoglobin in blood samples, there are multiple types of hemoglobin. In addition to normal hemoglobin (hemoglobin A), there exist multiple types of variant hemoglobin, also called abnormal hemoglobin, including hemoglobin C, hemoglobin D, hemoglobin E, hemoglobin S, etc. Electrophoresis (Patent Document 1) and liquid chromatography (Patent Document 2) are used for hemoglobin measurement.

[0003] When a blood sample containing abnormal hemoglobin E is measured by cation exchange chromatography, a hemoglobin E peak is detected between the stable hemoglobin A1c peak and the hemoglobin A0 peak. It has been reported that when a blood sample contains hemoglobin E, the (peak area of the) stable hemoglobin A1c peak decreases, and as a result, the measured stable hemoglobin A1c value is lower than the accurate stable hemoglobin A1c value. Therefore, to obtain an accurate stable hemoglobin A1c value, correction for increasing the hemoglobin A1c value is performed (Patent Documents 3 and 4).

[0004] Similarly, when measuring blood samples containing abnormal hemoglobin such as hemoglobin C, hemoglobin D, or hemoglobin S, correction is performed to obtain an accurate stable hemoglobin A1c value (Patent Document 5). In other words, in the technique described in Patent Document 5, when measuring the hemoglobin A1c value of a sample containing such abnormal hemoglobin, some of these abnormal hemoglobin components dissolve simultaneously with or after hemoglobin A0, which is the non-glycated component of hemoglobin A. Therefore, even if a prominent peak corresponding to the abnormal hemoglobin is excluded from the calculation of the stable hemoglobin A1c value, the stable hemoglobin A1c value will still show a low value. To address this phenomenon, a correction is performed to increase the hemoglobin A1c value based on the area value of the abnormal hemoglobin. [Prior art documents] [Patent Documents]

[0005] [Patent Document 1] Japanese Patent Publication No. 2019-078599 [Patent Document 2] Japanese Patent Application Publication No. 9-264889 [Patent Document 3] Japanese Patent Publication No. 2012-215470 [Patent Document 4] Japanese Patent Publication No. 2016-183871 [Patent Document 5] Japanese Patent Publication No. 2017-203677 [Overview of the project] [Problems that the invention aims to solve]

[0006] When measuring samples with known stable hemoglobin A1c values ​​using conventional measuring devices, it is often observed that the measured value is higher than the true stable hemoglobin A1c value. Our verification has revealed that when using samples that have deteriorated over time in separation and analysis methods based on cation exchange, the measured value of stable hemoglobin A1c increases.

[0007] Embodiments of this disclosure provide a measurement method that can measure a stable hemoglobin A1c value that is closer to the true value by removing as much as possible the effect of elevated stable hemoglobin A1c from the measured value of stable hemoglobin A1c, for example, when the sample has undergone deterioration such as degradation over time. [Means for solving the problem]

[0008] One aspect of the present disclosure is a method for measuring stable hemoglobin A1c by a hemoglobin separation analysis method based on the principle of cation exchange, comprising the steps of: obtaining an analytical signal from a blood sample to be measured by the separation analysis method; determining a C value, which is the peak value of the stable hemoglobin A1c peak, and an X value, which is the peak value of a specific peak appearing between the stable hemoglobin A1c peak and the hemoglobin A0 peak, from the analytical signal; and correcting the C value by applying the C value and X value to a predetermined calculation formula to obtain a C' value, which is a lower value of the C value, wherein the calculation formula is determined based on the correlation between the C value and X value obtained from a blood sample in which the stable hemoglobin A1c value is known by the separation analysis method. [Effects of the Invention]

[0009] According to embodiments of this disclosure, a measurement method is provided that can measure a stable hemoglobin A1c value that is closer to the true value by removing as much of the influence of the elevated stable hemoglobin A1c value as possible, even in cases where the sample has undergone deterioration such as degradation over time. [Brief explanation of the drawing]

[0010] [Figure 1] Electropherograms for sample 1, taken on the day of blood collection (A) and 28 days later (B). [Figure 2] A flowchart showing the first example of an arithmetic expression. [Figure 3] A flowchart showing a second example of an arithmetic expression. [Figure 4]Block diagram of the measuring device according to the embodiment. [Figure 5] Block diagram of the control unit. [Figure 6] Graphs showing the correlation between X values ​​and original C values ​​for each of the following samples: Sample 1 (A), Sample 2 (B), and Sample 3 (C). [Figure 7] A graph showing the correlation between the X value and the C' value for each of the following samples: Sample 1 (A), Sample 2 (B), and Sample 3 (C). [Figure 8] Graphs showing the change in C value over time for each of the following samples: Sample 1 (A), Sample 2 (B), and Sample 3 (C). [Figure 9] Graphs showing the time-dependent change in the rate of change of the C value relative to day 0 for each of the following samples: Sample 1 (A), Sample 2 (B), and Sample 3 (C). [Figure 10] A graph showing the correlation between the rate of change in the C value over time and the X value. [Figure 11] Graphs showing the change in C' value over time for each of the following samples: Sample 1 (A), Sample 2 (B), and Sample 3 (C). [Figure 12] Graphs showing the time-dependent change in the rate of change of the C' value relative to day 0 for each of the following samples: Sample 1 (A), Sample 2 (B), and Sample 3 (C). [Modes for carrying out the invention]

[0011] Hereinafter, embodiments of the present disclosure will be described with reference to the drawings. Common reference numerals in each drawing indicate the same parts unless otherwise specifically stated. In the present disclosure, the "peak value" refers to the height or area of each peak observed in an electropherogram as an analytical signal, and a relative value or an absolute value can be used. This relative value may be a ratio to the total area of the electropherogram, may be a ratio to the total peak area related to hemoglobin occupying the electropherogram, or may be a ratio to the area of a specific peak (e.g., hemoglobin A0 peak). The analytical signal may be a chromatogram as well as an electropherogram. The meaning of "peak value" in that case is the same as described above.

[0012] In the following description, hemoglobin A0 is denoted as "HbA0" and stable hemoglobin A1c is denoted as "s-HbA1c". Further, in an electropherogram of hemoglobin observed when a human-derived blood sample is subjected to capillary electrophoresis, peaks attributable to HbA0 and s-HbA1c are referred to as "HbA0 peak" and "s-HbA1c peak", respectively. In addition, peak values for the HbA0 peak, the s-HbA1c peak, and a "specific peak" described later are referred to as "HbA0 peak value", "s-HbA1c peak value", and "specific peak value", respectively.

[0013] Figure 1 shows an electropherogram (A) of hemoglobin measured on the day of blood collection by capillary electrophoresis based on cation exchange principle for Sample 1 among blood samples of Examples described later, and an electropherogram (B) of hemoglobin measured in the same manner 28 days after blood collection. In FIG. 1(A) and FIG. 1(B), an HbA0 peak 50 around an elution time of 27 seconds and an s-HbA1c peak 60 around an elution time of 20 seconds are observed.

[0014] In Figure 1(B), a peak not observed in Figure 1(A) appears around 22.5 seconds, at the elution time between the s-HbA1c peak 60 and the HbA0 peak 50. This peak is referred to as the "specific peak 70". The reason for the appearance of specific peak 70 is unknown, but it may appear in altered blood samples, such as blood samples that have deteriorated over time, having passed several days since collection. When the detection time of the peak top of the s-HbA1c peak 60 is set to 0 and the detection time of the peak top of the HbA0 peak 50 is set to 1, specific peak 70 is a peak that appears at a detection time greater than 0 but less than 1, preferably between 0.01 and 0.8, more preferably between 0.1 and 0.65, and even more preferably between 0.2 and 0.5 (0.41 in Figure 1(B)).

[0015] As shown in the examples described later, when hemoglobin in blood samples stored for various days is measured using capillary electrophoresis, which is based on the principle of cation exchange, a positive correlation is observed between the s-HbA1c peak value and a specific peak value.

[0016] As described above, the s-HbA1c measurement method of this embodiment is based on a hemoglobin separation and analysis method that uses cation exchange as its principle, and includes the steps of: obtaining an analytical signal from a blood sample to be measured by the separation and analysis method; determining the C value, which is the peak value of the s-HbA1c peak, and the X value, which is the peak value of a specific peak that appears between the s-HbA1c and HbA0 peaks, from the analytical signal; and correcting the C value by applying the C value and X value to a predetermined calculation formula to obtain a C' value which is a lower value of the C value. This calculation formula is determined based on the correlation between the C value and X value obtained from a blood sample whose s-HbA1c value is known by the separation and analysis method.

[0017] The calculation formula may include, for example, a calculation step of obtaining a Y value by multiplying the X value obtained from the blood sample to be measured by a Y value, which is the slope of the regression line between the C value and X value obtained from multiple blood samples with known s-HbA1c values, and a calculation step of obtaining a C' value by subtracting the Y value from the C value obtained from the blood sample to be measured. Alternatively, a regression line between the C value and X value may be obtained for each of the multiple blood samples, and the average of their slopes may be used as the A value.

[0018] In other words, the C value and X value are measured for each of several blood samples with known s-HbA1c values, and the A value is determined in advance as the slope of the regression line between the multiple C values ​​and the corresponding multiple X values. Then, for the blood sample to be measured, an analytical signal is obtained using the separation analysis method described above, and the C value and X value are measured. These C value and X value are applied to the calculation formula shown in the flowchart in Figure 2, for example.

[0019] Specifically, in the step shown in S10, the Y value is obtained by multiplying the X value measured in the blood sample to be measured by the A value, as shown in formula (1) below.

[0020] Y = A * X ... (1)

[0021] This Y value is considered to be the value that corresponds to the amount by which the C value is raised relative to the true value in accordance with the X value. Next, in the step shown in S15, as shown in formula (2) below, the C value is reduced by obtaining the C value obtained by subtracting the Y value from the C value measured in the blood sample to be measured.

[0022] C' = CY ···(2)

[0023] Note that the Y value obtained by formula (1) above may overestimate or underestimate the amount that the C value has boosted relative to the true value. In such cases, the Y value may be corrected to an appropriate value by modifying the A value in formula (1) above, modifying the X value, adding or subtracting an arbitrary value to the product of the A value and the X value, or by using two or more of these methods in combination.

[0024] In addition to the above, the calculation formula may also include, for example, a calculation step of obtaining the R1 value, which is the rate of change of the C value, by applying the X value obtained from the blood sample to be measured to the correlation between the rate of change of the C value over time obtained from a blood sample with a known s-HbA1c value and the X value, and a calculation step of obtaining the C' value by subtracting the value obtained by multiplying the C value obtained from the blood sample to be measured by the R1 value from the C value.

[0025] In other words, the C value and X value are measured for each of several blood samples with known s-HbA1c values. At this time, the C value and X value are measured for each of the same blood samples from the day of collection to after a predetermined number of days of storage. The correlation between the rate of change over time, which is the rate at which the C value changes after a predetermined number of days of storage relative to the C value on the day of collection, and the corresponding multiple X values, for example, a regression line, is determined in advance. If the rate of change over time is y and the X value is x, then this regression line is given by equation (3) below.

[0026] y = ax + b ... (3)

[0027] Here, in equation (3) above, a is the slope of the regression line and b is the intercept of the regression line. Then, an analytical signal is obtained by the separation and analysis method described above, and the C value and X value are measured for the blood sample to be measured. These C value and X value are applied to the calculation formula shown in the flowchart in Figure 3.

[0028] Specifically, in the step shown in S20, the value of y obtained by substituting the X value for x in equation (3) above, as shown in equation (4) below, becomes the R1 value.

[0029] R1 = a*X + b ... (4)

[0030] This R1 value is the rate of change over time of the C value, which corresponds to the X value. Next, in the step shown in S25, as shown in equation (5) below, the C value is reduced by subtracting the value obtained by multiplying the C value measured in the blood sample to be measured by this R1 value.

[0031] C′=CC*R1···(5)

[0032] Note that the R1 value obtained by formula (4) above may result in the C value being excessively underestimated or underestimated in the correction performed by formula (5) above. In such cases, the R1 value may be corrected to an appropriate value by modifying the value of a in formula (4), modifying the value of b, modifying the X value, or using two or more of these in combination.

[0033] The calculation formula may include a correlation table determined based on the correlation between the C value and X value obtained from a blood sample whose s-HbA1c value is known by the separation analysis method. For example, the C' value may be obtained by using the correlation table to determine a value or percentage equivalent to the boost from the X value and C value measured in the blood sample to be measured, and then subtracting that value from the C value. Alternatively, the Y value in formula (1) may be obtained using the correlation table from the X value and C value measured in the blood sample to be measured, and then subtracting the Y value from the C value to obtain the C' value. Alternatively, the R1 value in formula (4) may be obtained based on the correlation table from the X value and C value measured in the blood sample to be measured, and then subtracting the value obtained by multiplying the C value by the R1 value from the C value to obtain the C' value.

[0034] Here, the specific peak is preferably a peak that appears when the detection time of the peak top of the s-HbA1c peak is set to 0 and the detection time of the peak top of the HbA0 peak is set to 1, with the detection time being greater than 0 and less than 1, preferably between 0.01 and 0.8, more preferably between 0.1 and 0.65, and even more preferably between 0.2 and 0.5.

[0035] For example, in Figure 1(B), if we set the time at which the peak top of the s-HbA1c peak 60 is detected, which is an elution time of 20 seconds, to 0, and the time at which the peak top of the HbA0 peak 50 is detected, which is an elution time of 26.6 seconds, to 1, then the time at which the peak top of the specific peak 70 is detected, which is an elution time of 22.7 seconds, will be 0.41.

[0036] An increase in the C value is a phenomenon observed when a specific peak appears. In other words, as the X value, which is the peak value of the specific peak, increases, the C value is raised and becomes higher. The reason why a specific peak appears is unknown, but it may appear in deteriorated blood samples, such as blood samples that have deteriorated over time since the day of collection. Therefore, from the perspective of not unnecessarily correcting the C value for blood samples that have not deteriorated, a threshold for the X value may be set to determine whether or not to perform correction. Then, if the X value exceeds the threshold (for example, 5% of total hemoglobin), the C value may be corrected using the above calculation formula.

[0037] As shown in Figure 4, the s-HbA1c measuring device 10 of this embodiment includes a storage device 150 that stores a predetermined calculation formula based on the correlation between the C value, which is the peak value of the s-HbA1c peak, and the X value, which is the peak value of a specific peak that appears between the s-HbA1c peak and the HbA0 peak, when a blood sample with a known s-HbA1c value is subjected to a hemoglobin separation and analysis method based on the principle of cation exchange; an analyzer 20 that obtains an analysis signal by subjecting the blood sample to be measured to a hemoglobin separation and analysis method based on the principle of cation exchange; a first calculation device 160 that obtains the C value, which is the peak value of the s-HbA1c peak, and the X value, which is the peak value of a specific peak that appears between the s-HbA1c peak and the HbA0 peak, from the analysis signal; and a second calculation device 170 that calculates a C' value by applying the C value and X value to the calculation formula and lowering the C value.

[0038] The analyzer 20 is a device that performs separation and analysis of hemoglobin based on the principle of cation exchange. For example, a capillary electrophoresis apparatus or a liquid chromatography apparatus can be used as the analyzer 20. The analyzer 20 separates hemoglobin into its components by cation exchange and outputs signal data corresponding to each component, for example, as an electropherogram (in the case of a capillary electrophoresis apparatus) or a chromatogram (in the case of a liquid chromatography apparatus).

[0039] The control device 100 includes a storage device 150, a first arithmetic unit 160, and a second arithmetic unit 170. It controls the analysis device 20 and performs various calculations based on the signal data output from the analysis device 20.

[0040] As shown in the hardware configuration of Figure 5, the control device 100 includes a CPU (Central Processing Unit) 110, a ROM (Read Only Memory) 120, a RAM (Random Access Memory) 130, and a storage device 150. Each component is connected to the others via a bus 190 so that they can communicate with each other.

[0041] The CPU 110 is a central processing unit that executes various programs and controls various parts. Specifically, the CPU 110 reads a program from the ROM 120 or storage device 150 and executes the program using the RAM 130 as a working area. The CPU 110 controls the analysis device 20 according to the program recorded in the ROM 120 or storage device 150.

[0042] ROM 120 stores various programs and data. RAM 130 temporarily stores programs or data as a working area. The storage device 150 is configured as storage using an HDD (Hard Disk Drive), SSD (Solid State Drive), or flash memory, and stores various programs, including the operating system, and various data. In this embodiment, ROM 120 or storage device 150 stores programs and various data related to control and calculations. This various data also includes the calculation formulas described above.

[0043] The control device 100, with its CPU 110, executes the program described above. First, as the first arithmetic unit 160, it obtains the C value and X value from the analysis signal from the analysis device 20. Then, as the second arithmetic unit 170, it applies the C value and X value to the arithmetic formula stored in the storage device 150 to calculate the C' value, which is a lower value of the C value.

[0044] The s-HbA1c measurement process, which is read and executed by the CPU 110, may also be performed by various processors other than the CPU 110. Examples of such processors include dedicated electrical circuits, which are processors with circuit configurations specifically designed to perform particular processing, such as FPGAs (Field-Programmable Gate Arrays), PLDs (Programmable Logic Devices) whose circuit configurations can be changed after manufacturing, and ASICs (Application Specific Integrated Circuits). Furthermore, the s-HbA1c measurement process may be performed by one of these various processors, or by a combination of two or more processors of the same or different types (for example, multiple FPGAs or a combination of a CPU and an FPGA). More specifically, the hardware structure of these various processors is an electrical circuit that combines circuit elements such as semiconductor elements. [Examples]

[0045] (1) Measurement data For each of the three subjects, venous blood was collected into nine different blood collection tubes containing anticoagulants (see Table 1 below) to obtain whole blood samples (Sample 1, Sample 2, and Sample 3). On the day of collection, these samples were subjected to dual C and X values ​​on day 0 using a capillary electrophoresis apparatus (The Lab 001, Arkray). Subsequently, the samples were stored at -20°C, and dual C and X values ​​were measured at 1 day, 3 days, 7 days, 14 days, and 28 days after storage.

[0046] [Table 1]

[0047] (2) First correction method Figure 6 plots data from 108 sets of samples (1A, 2B, and 3C) for each of the three samples, obtained by taking two measurements at six test points using nine different blood collection tubes. The x-axis represents the X-value, and the y-axis represents the raw measured C-value (original C-value, in units of ‰). The dashed lines in each graph represent the regression lines between the X-value and the original C-value. The slopes of the regression lines were 0.382 for sample 1, 0.348 for sample 2, and 0.339 for sample 3. The correlation coefficients of the regression lines were 0.744 for sample 1, 0.715 for sample 2, and 0.637 for sample 3, all showing a positive correlation.

[0048] Then, the average value of the slopes mentioned above, 0.356, is taken as A in Figure 2. This value is multiplied by the X value obtained from the blood sample measured in Figure 6 to obtain the Y value. The C' value, which is corrected by subtracting this Y value from the C value obtained from the same blood sample to reduce its value, is plotted in a graph in the same way as in Figure 6, corresponding to the X value, as shown in Figure 7. The dashed lines in each graph are the regression lines between the X value and the C' value. The slopes of the regression lines were -0.072 for sample 1, -0.103 for sample 2, and -0.106 for sample 3. The correlation coefficients of the regression lines were -0.200 for sample 1, -0.276 for sample 2, and -0.252 for sample 3.

[0049] The uncorrected and corrected data for each of the samples 1 to 3 were converted from raw values ​​to internationally standardized values, and the mean, standard deviation, and coefficient of variation are shown in Table 2 below.

[0050] [Table 2]

[0051] Table 2 above shows that correcting from the original C value to the C' value equalizes the variability in the data and suppresses the increase in C value that occurs due to the increase in X value caused by sample storage.

[0052] (3)Second correction method Figure 8 is a graph plotted for each blood collection tube, using the same measurement data for each of the samples 1(A), 2(B), and 3(C) as in the first correction method described above. The average value obtained from duplicate measurements at each measurement point is plotted with the number of storage days on the horizontal axis and the value converted from the raw measured C value to the internationally standardized value (in %) on the vertical axis. Figure 9 is a graph plotted with the rate of change over time (rate of change over time, in %) calculated from each C value in Figure 8 relative to the C value on day 0, plotted on the vertical axis and the number of storage days on the horizontal axis. Here, the rate of change over time (R2) is calculated by dividing the C value on day 0 by C0 and the C value on day n by C n Therefore, the value is calculated using the following formula (6).

[0053] R2=(C n -C0) / C0*100 ···(6)

[0054] In all samples, regardless of the type of blood collection tube used, the C value (Figure 8) increased as the storage period progressed, and consequently, the rate of change in the C value over time (Figure 9) also tended to increase.

[0055] Figure 10 is a scatter plot showing the correlation between the time-dependent rate of change of the C value and the X value for each measurement point. From this scatter plot, it can be seen that there is a positive correlation between the rate of change of the X value and the rate of change of the C value. The dashed line in the figure represents the regression line, and if the time-dependent rate of change is y and the X value is x, it is expressed by the following equation (7). The correlation coefficient (r) of this regression line was 0.782.

[0056] y = 0.6323x - 1.4143 ... (7)

[0057] Then, the X value obtained from the blood sample to be measured is set as x, and the value of y obtained by substituting it into the above formula (7) is taken as the R1 value. The C value is corrected by subtracting the value obtained by multiplying this R1 value by the C value to make it lower, and the C' value is plotted on the horizontal axis with the number of storage days, similar to Figure 8, as shown in Figure 11. Furthermore, Figure 12 plots the rate of change over time relative to the C' value on day 0, calculated from each C' value in Figure 11, on the vertical axis and the number of storage days on the horizontal axis.

[0058] In all samples, regardless of the type of blood collection tube used, the increase in C' value (Figure 11) was suppressed even after the storage period, compared to Figure 8, and the rate of change in C' value over time (Figure 12) was also leveled out.

[0059] Table 3 below shows the mean, standard deviation, and coefficient of variation of each data set for Sample 1 to Sample 3, both before and after correction.

[0060] [Table 3]

[0061] From Table 3 above, it can be seen that the coefficient of variation is smaller after correction compared to before correction, indicating that the accuracy of each measurement has improved. [Industrial applicability]

[0062] This invention can be used to measure stable HbA1c in blood samples using a hemoglobin separation and analysis method based on the principle of cation exchange. [Explanation of symbols]

[0063] 10 Measuring device 20 Analyzer 50 HbA0 peak 60 s-HbA1c peak 70 Specific Peak 100 Control device 110 CPU 120 ROM 130 RAM 150 Storage device 160 1st calculation unit 170 Second computing unit 190 bus

Claims

1. A method for measuring stable hemoglobin A1c by a hemoglobin separation and analysis method based on the principle of cation exchange, A step of obtaining an analytical signal from a blood sample to be measured by the aforementioned separation and analysis method, A step of determining the C value, which is the peak value of the stable hemoglobin A1c peak, and the X value, which is the peak value of a specific peak appearing between the stable hemoglobin A1c peak and the hemoglobin A0 peak, from the aforementioned analysis signal, and A step of correcting the C value by applying the C value and the X value to a predetermined calculation formula, and subtracting from the C value a value equivalent to the amount by which the C value has been raised relative to the true value according to the X value, thereby obtaining a C' value which is a lower value of the C value. Includes, The aforementioned calculation formula is determined based on the correlation between the C value and X value obtained from a blood sample whose stable hemoglobin A1c value is known by the aforementioned separation and analysis method. Method for measuring stable hemoglobin A1c.

2. The aforementioned calculation formula is, Y = A * X And, C' = C - Y A method for measuring stable hemoglobin A1c according to claim 1, comprising the formula represented by (wherein A is the slope of the regression line between the X value obtained from a plurality of blood samples with known stable hemoglobin A1c values ​​and the C value obtained from a plurality of blood samples with known stable hemoglobin A1c values, X is the X value obtained from the blood sample to be measured, C is the C value obtained from the blood sample to be measured, and Y is a value corresponding to the amount by which the C value is raised relative to the true value according to the X value).

3. The aforementioned calculation formula is, R １ =a*X+b and C′=C-C*R １ A method for measuring stable hemoglobin A1c according to claim 1, comprising the formula represented by (wherein a is the slope of the regression line between the rate of change over time of the X value obtained from a plurality of blood samples with known stable hemoglobin A1c values ​​and the C value obtained from a plurality of blood samples with known stable hemoglobin A1c values, b is the intercept of the regression line, X is the X value obtained from the blood sample to be measured, C is the C value obtained from the blood sample to be measured, and R1 is the rate of change over time of the C value corresponding to the X value).

4. A method for measuring stable hemoglobin A1c according to any one of claims 1 to 3, wherein when the peak top detection time of the stable hemoglobin A1c peak is set to 0 and the peak top detection time of the hemoglobin A0 peak is set to 1, the peak that appears at a detection time of 0.1 or more and less than 0.65 is defined as the specific peak.

5. The method for measuring stable hemoglobin A1c according to any one of claims 1 to 3, wherein the correction step is performed when the X value exceeds a threshold.

6. A storage device that stores a predetermined calculation formula based on the correlation between the C value, which is the peak value of the stable hemoglobin A1c peak, and the X value, which is the peak value of a specific peak that appears between the stable hemoglobin A1c peak and the hemoglobin A0 peak, indicating alteration of the blood sample, when a blood sample with a known stable hemoglobin A1c value is subjected to a hemoglobin separation and analysis method based on the principle of cation exchange. The blood sample to be measured is subjected to a hemoglobin separation and analysis method based on the principle of cation exchange. Analytical device for obtaining analytical signals, A first calculation device that obtains from the aforementioned analysis signal the C value, which is the peak value of the stable hemoglobin A1c peak, and the X value, which is the peak value of a specific peak that appears between the stable hemoglobin A1c peak and the hemoglobin A0 peak, associated with the alteration of the blood sample, and A second calculation device calculates a C' value obtained by applying the C value and the X value to the calculation formula and subtracting from the C value a value corresponding to the amount by which the C value has been raised relative to the true value according to the X value, thereby lowering the C value. A device for measuring stable hemoglobin A1c, comprising the above.

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