A multi-mode hemoglobin eluate and a method for measuring the same

Abstract

The present invention discloses a multi-mode hemoglobin eluent and a method for measuring the same. The eluent includes eluent A, eluent B, eluent C, eluent E, eluent F, and eluent G for measuring hemoglobin. The raw materials of eluent A include 0.08-0.12 wt% of citric acid monohydrate, 0.40-0.50 wt% of tri-alkali metal citrate hydrate, 0.3-0.5 wt% of alkali metal perchlorate, and 0.01-0.04 wt% of sodium hydroxide. The present invention achieves the separation and measurement of multiple hemoglobin modes using a single eluent without requiring replacement of a chromatographic column stationary phase or mobile phase. The method can adapt to different hemoglobin measurement requirements, including conventional hemoglobin measurement, mutant hemoglobin measurement, and thalassemia-related hemoglobin measurement.

Description

Technical Field

[0001] The present invention relates to the technical field of hemoglobin determination, and in particular to a multi-mode hemoglobin determination eluate and a determination method thereof. Background Art

[0002] Hemoglobins, particularly glycated hemoglobin (HbA1c), a glycosylated protein formed by the binding of hemoglobin with glucose, reflect average blood sugar levels over the past one to two months. Therefore, they are widely used for testing for lifestyle-related diseases, including diabetes and metabolic syndrome, and for blood sugar management. Therefore, methods for easily and accurately measuring hemoglobins such as HbA1c are in high demand.

[0003] In the prior art, methods for measuring HbA1c include high-performance liquid chromatography (HPLC), immunoassays, enzymatic methods, and electrophoresis. HPLC is the standard method for measuring HbA1c and is widely used in clinical testing. HPLC methods for separating and measuring hemoglobins generally use a cationic column as the stationary phase and a phosphate- or succinate-based buffer as the eluent. This method is generally effective in separating and measuring HbA1a, HbA1b, HbF, L-A1c, HbA1c, and HbA0. Separating and measuring hemoglobins other than those listed above, such as HbA2, HbE, HbD, HbS, and HbC, requires switching between different columns and eluents. This is time-consuming and inconvenient in terms of both operation and performance, not only inefficient but also impacting measurement accuracy and reproducibility. Examples include Bio-Rad and ARKRAY. Summary of the Invention

[0004] In response to the problems existing in the prior art, the present invention provides a multi-mode hemoglobin determination eluent and a determination method thereof. The present invention realizes the separation and determination modes of multiple hemoglobins using one eluent without replacing the chromatographic column stationary phase and mobile phase.

[0005] In order to achieve the above object, the present invention provides a multi-mode hemoglobin determination eluent, wherein the eluent includes eluent A, eluent B, eluent C, eluent E, eluent F and eluent G for determining hemoglobin;

[0006] The raw materials of the eluent A include 0.08wt%-0.12wt% of citric acid monohydrate, 0.40wt%-0.50wt% of tri-alkali metal citrate hydrate, 0.3wt%-0.5wt% of alkali metal perchlorate and 0.01wt%-0.04wt% of sodium hydroxide;

[0007] The raw materials of the eluent B include 0.01wt%-0.08wt% of citric acid monohydrate, 0.50wt%-0.60wt% of tri-alkali metal citrate hydrate, 1.0wt%-2.0wt% of alkali metal perchlorate and 0.02wt%-0.08wt% of sodium hydroxide;

[0008] The raw materials of the eluent C include 0.001wt%-0.01wt% of citric acid monohydrate, 0.50wt%-0.60wt% of tri-alkali metal citrate hydrate, 0.05wt%-0.15wt% of alkali metal perchlorate and 0.02wt%-0.08wt% of sodium hydroxide;

[0009] The eluent E is a mixture of eluent A and eluent C, with a weight ratio of 0.4-1.0;

[0010] The eluent F is a mixture of eluent B and eluent C, with a weight ratio of 0.1 to 0.8;

[0011] The eluent G is prepared by mixing the eluent C and the eluent A, with the weight ratio being 0.25 to 0.5.

[0012] Preferably, the osmotic pressure of the eluent A is 100 mOsm-200 mOsm, the pH value is 5.0-6.0, and the conductivity is 8.2 sm / cm-8.6 sm / cm;

[0013] The eluent B has an osmotic pressure of 500 mOsm-700 mOsm, a pH of 6.0-7.0, and a conductivity of 25 sm / cm-35 sm / cm;

[0014] The eluent C has an osmotic pressure of 10 mOsm-200 mOsm, a pH of 6.0-7.0, and a conductivity of 3.0 sm / cm to 6.0 sm / cm.

[0015] Preferably, the hemoglobin is selected from one or more of HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0, HbA2, HbE, HbD, HbS and HbC.

[0016] The present invention also provides a method for measuring hemoglobin eluate using the multi-mode assay, comprising the following steps:

[0017] Step X1: Select the corresponding eluent, eluent combination, and measurement mode according to the type of hemoglobin to be measured;

[0018] Step X2: injecting a filler into a high-pressure liquid delivery pump and delivering the selected eluent or eluent combination into the chromatographic column using the high-pressure liquid delivery pump;

[0019] Step X3: separating and measuring hemoglobin in a chromatographic column;

[0020] Preferably, the filler is a methacrylate copolymer, which is used to connect the high-pressure liquid delivery pump and the eluent and to separate and measure hemoglobin; the cation exchange group of the chromatographic column is one or more of sulfonyl, carboxyl and phosphate groups.

[0021] Preferably, the assay modes include rapid mode, rapid variation mode, variation assay mode and thalassemia mode;

[0022] Among them, the rapid mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c and HbA0 hemoglobin;

[0023] The rapid mutation mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0 hemoglobins, and mutant hemoglobins V-Win, where mutant hemoglobins V-Win are a general term for HbE, HbD, HbS, and HbC;

[0024] The variation pattern is used to separate the determination of HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0 hemoglobin and HbE, HbD, HbS and HbC;

[0025] The thalassemia mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0, HbA2, HbE, HbD, HbS and HbC.

[0026] Preferably, the specific steps in the fast mode are:

[0027] S1: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 2.0 mL / min and a delivery time of 25 s;

[0028] S2: Use a high-pressure liquid delivery pump to deliver eluent B to the separation column at a flow rate of 2.0 mL / min and a delivery time of 10 s;

[0029] S3: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 20 s.

[0030] Preferably, the specific steps in the rapid mutation mode are:

[0031] S4: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 30 s;

[0032] S5: Use a high-pressure liquid delivery pump to deliver eluent C to the separation column at a flow rate of 1.8 mL / min and a delivery time of 25 s.

[0033] S6: Use a high-pressure liquid delivery pump to deliver eluent B to the separation column at a flow rate of 1.8 mL / min and a delivery time of 15 s.

[0034] S7: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 30 s.

[0035] Preferably, the specific steps in the mutation mode are:

[0036] S8: Use a high-pressure liquid delivery pump to deliver eluent A to the separation column: flow rate is 1.8 mL / min, delivery time is 35 s;

[0037] S9: Use a high-pressure liquid delivery pump to deliver eluent C to the separation column at a flow rate of 1.8 mL / min and a delivery time of 45 s.

[0038] S10: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 10 s;

[0039] S11: Use a high-pressure liquid delivery pump to deliver eluent C to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 10 s.

[0040] S12: Use a high-pressure liquid delivery pump to deliver eluent B to the separation column at a flow rate of 1.8 mL / min and a delivery time of 10 s.

[0041] S13: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 30 seconds.

[0042] Preferably, the specific steps in the thalassemia model are:

[0043] S14: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 35 s;

[0044] S15: Use a high-pressure liquid delivery pump to deliver eluent C to the separation column at a flow rate of 1.8 mL / min and a delivery time of 25 s.

[0045] S16: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 10 s;

[0046] S17: Use a high-pressure liquid delivery pump to deliver eluent E to the separation chromatographic column, and continuously switch and mix the liquids through a high-frequency solenoid valve at a flow rate of 1.8 mL / min for 120 s.

[0047] S18: Use a high-pressure liquid delivery pump to deliver eluent F to the separation chromatographic column, and continuously switch and mix the liquids through a high-frequency solenoid valve at a flow rate of 1.8 mL / min for 100 s.

[0048] S19: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 50 s.

[0049] The technical solution of the present invention has the following beneficial effects:

[0050] The eluent of the present invention eliminates the need to replace the eluent when separating and measuring different hemoglobin types. Simply switching the fluid circuit timing control allows for free combination, mixing, and delivery of the eluents, achieving the same effect as replacing the reagents. This system can meet the rapid testing needs of separating and measuring only HbA1a, HbA1b, HbF, L-A1c, HbA1c, and HbA0 hemoglobin, while also meeting the needs of separating and measuring HbA2, HbE, HbD, HbS, and HbC.

[0051] The present invention has multimodal applicability: by providing a variety of eluents and measurement modes, the present invention can adapt to different types of hemoglobin measurement needs, including conventional hemoglobin measurement, mutant hemoglobin measurement and thalassemia-related hemoglobin measurement.

[0052] The method of the present invention simplifies the operation process, does not need to replace the chromatographic column and eluent, and can achieve efficient separation and determination of different hemoglobins by simply changing the combination and ratio of the eluent.

[0053] The present invention is highly time-efficient: by eliminating the steps of replacing the chromatographic column and eluent, the method of the present invention greatly shortens the measurement time and improves work efficiency, making it particularly suitable for emergency and batch testing needs; the simplified operating process and reduced reagent consumption reduce the overall operating cost, making the method of the present invention more economical and affordable.

[0054] The eluent composition of the present invention is carefully designed to ensure the accuracy and repeatability of the measurement results and improve the reliability of the detection.

[0055] The eluent composition of the present invention is suitable for the determination of various hemoglobins, including but not limited to HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0, HbA2, HbE, HbD, HbS and HbC, making it a powerful tool for hemoglobin research and clinical diagnosis. DETAILED DESCRIPTION

[0056] The present invention is further described below with reference to specific embodiments.

[0057] The present invention provides a multi-mode hemoglobin determination eluent, wherein the eluent includes eluent A, eluent B, eluent C, eluent E, eluent F and eluent G for determining hemoglobin;

[0058] The raw materials of the eluent A include 0.08wt%-0.12wt% of citric acid monohydrate, 0.40wt%-0.50wt% of tri-alkali metal citrate hydrate, 0.3wt%-0.5wt% of alkali metal perchlorate and 0.01wt%-0.04wt% of sodium hydroxide;

[0059] The raw materials of the eluent B include 0.01wt%-0.08wt% of citric acid monohydrate, 0.50wt%-0.60wt% of tri-alkali metal citrate hydrate, 1.0wt%-2.0wt% of alkali metal perchlorate and 0.02wt%-0.08wt% of sodium hydroxide;

[0060] The raw materials of the eluent C include 0.001wt%-0.01wt% of citric acid monohydrate, 0.50wt%-0.60wt% of tri-alkali metal citrate hydrate, 0.05wt%-0.15wt% of alkali metal perchlorate and 0.02wt%-0.08wt% of sodium hydroxide;

[0061] The eluent E is a mixture of eluent A and eluent C, with a weight ratio of 0.4-1.0;

[0062] The eluent F is a mixture of eluent B and eluent C, with a weight ratio of 0.1 to 0.8;

[0063] The eluent G is prepared by mixing the eluent C and the eluent A, with the weight ratio being 0.25 to 0.5.

[0064] Preferably, the osmotic pressure of the eluent A is 100 mOsm-200 mOsm, the pH value is 5.0-6.0, and the conductivity is 8.2 sm / cm-8.6 sm / cm;

[0065] Osmolality (100 mOsm-200 mOsm): This range helps maintain the natural state of cells in the sample, reduces hemolysis, and ensures mild conditions during the elution process to avoid denaturation or degradation of hemoglobin; pH (5.0-6.0): This pH range provides a stable environment for hemoglobin, preventing it from changing under acidic conditions, while providing suitable pH conditions for chromatographic analysis; Conductivity (8.2 sm / cm-8.6 sm / cm): This conductivity range helps control the migration speed of ions in the eluent, optimize separation efficiency, shorten analysis time, and ensure sufficient resolution.

[0066] The eluent B has an osmotic pressure of 500 mOsm-700 mOsm, a pH of 6.0-7.0, and a conductivity of 25 sm / cm-35 sm / cm;

[0067] Osmolarity (500 mOsm-700 mOsm): Higher osmolarity allows for more efficient separations in certain chromatographic analyses, particularly when analyzing larger molecules or hemoglobin variants that require higher elution strengths. pH (6.0-7.0): A near-neutral pH range helps preserve the native structure of hemoglobin, preventing denaturation that can occur under alkaline conditions and ensuring assay accuracy.

[0068] Conductivity (25 sm / cm-35 sm / cm): Higher conductivity helps accelerate ion migration and improve separation efficiency, making it suitable for applications requiring rapid elution.

[0069] The eluent C has an osmotic pressure of 10 mOsm-200 mOsm, a pH of 6.0-7.0, and a conductivity of 3.0 sm / cm to 6.0 sm / cm.

[0070] Conductivity (3.0 sm / cm to 6.0 sm / cm): This moderate conductivity range allows for faster separations while maintaining adequate resolution, making it suitable for applications that require a balance between analysis time and resolution.

[0071] Furthermore, the hemoglobin level is selected from one or more of HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0, HbA2, HbE, HbD, HbS, and HbC. By measuring multiple hemoglobin variants, more comprehensive diagnostic information for blood disorders can be provided, such as diabetes control (via HbA1c), thalassemia (via HbA2), and other inherited hemoglobinopathies (such as HbE and HbS). Early diagnosis: Certain hemoglobin variants, such as HbA1c, are indicators of long-term blood sugar control and facilitate early detection of diabetes-related complications. Regular monitoring of specific hemoglobin variant levels is crucial for assessing treatment effectiveness and adjusting treatment plans.

[0072] The present invention also provides a method for measuring hemoglobin eluate using the multi-mode assay, comprising the following steps:

[0073] Step X1: Select the corresponding eluent, eluent combination, and measurement mode according to the type of hemoglobin to be measured;

[0074] Step X2: injecting a filler into a high-pressure liquid delivery pump and delivering the selected eluent or eluent combination into the chromatographic column using the high-pressure liquid delivery pump;

[0075] Step X3: separating and measuring hemoglobin in a chromatographic column;

[0076] Preferably, the filler is a methacrylate copolymer, which is used to connect the high-pressure liquid delivery pump and the eluent and to separate and measure hemoglobin; the cation exchange group of the chromatographic column is one or more of sulfonyl, carboxyl and phosphate groups.

[0077] Preferably, the assay modes include rapid mode, rapid variation mode, variation assay mode and thalassemia mode;

[0078] Among them, the rapid mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c and HbA0 hemoglobin; it is easy to operate, takes little time, and is suitable for rapid screening of a large number of samples. Application: Commonly used for routine physical examinations and rapid testing of outpatients.

[0079] The rapid mutation mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0 hemoglobins, and mutant hemoglobins V-Win, where mutant hemoglobins V-Win are a general term for HbE, HbD, HbS, and HbC;

[0080] The rapid mutation mode adds the detection of mutant hemoglobin on the basis of the rapid mode, expands the detection range, and is suitable for the preliminary screening and diagnosis of hereditary hemoglobin diseases.

[0081] The variant mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0 hemoglobins and HbE, HbD, HbS and HbC; it provides more precise separation and measurement, capable of detecting more types of variant hemoglobins, and is suitable for the diagnosis and typing of patients suspected of hereditary hemoglobinopathies.

[0082] The thalassemia mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0, HbA2, HbE, HbD, HbS and HbC. It can comprehensively cover the hemoglobin types related to thalassemia and provide detailed measurement results. It is suitable for screening, diagnosis and treatment monitoring of thalassemia.

[0083] The present invention can adapt to different detection needs by providing multiple measurement modes. Whether it is rapid screening or detailed diagnosis, the rapid mode and rapid mutation mode greatly shorten the measurement time and improve the detection efficiency. The mutation mode and thalassemia mode provide more precise separation and measurement, ensuring the accuracy of the results; the unified measurement process and automated operation reduce the complexity of the operation and lower the skill requirements of the operator.

[0084] Preferably, the specific steps in the fast mode are:

[0085] S1: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 2.0 mL / min and a delivery time of 25 s;

[0086] S2: Use a high-pressure liquid delivery pump to deliver eluent B to the separation column at a flow rate of 2.0 mL / min and a delivery time of 10 s;

[0087] S3: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 20 s.

[0088] Preferably, the specific steps in the rapid mutation mode are:

[0089] S4: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 30 s;

[0090] S5: Use a high-pressure liquid delivery pump to deliver eluent C to the separation column at a flow rate of 1.8 mL / min and a delivery time of 25 s.

[0091] S6: Use a high-pressure liquid delivery pump to deliver eluent B to the separation column at a flow rate of 1.8 mL / min and a delivery time of 15 s.

[0092] S7: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 30 s.

[0093] Preferably, the specific steps in the mutation mode are:

[0094] S8: Use a high-pressure liquid delivery pump to deliver eluent A to the separation column: flow rate is 1.8 mL / min, delivery time is 35 s;

[0095] S9: Use a high-pressure liquid delivery pump to deliver eluent C to the separation column at a flow rate of 1.8 mL / min and a delivery time of 45 s.

[0096] S10: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 10 s;

[0097] S11: Use a high-pressure liquid delivery pump to deliver eluent C to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 10 s.

[0098] S12: Use a high-pressure liquid delivery pump to deliver eluent B to the separation column at a flow rate of 1.8 mL / min and a delivery time of 10 s.

[0099] S13: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 30 seconds.

[0100] Preferably, the specific steps in the thalassemia model are:

[0101] S14: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 35 s;

[0102] S15: Use a high-pressure liquid delivery pump to deliver eluent C to the separation column at a flow rate of 1.8 mL / min and a delivery time of 25 s.

[0103] S16: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 10 s;

[0104] S17: Use a high-pressure liquid delivery pump to deliver eluent E to the separation chromatographic column, and continuously switch and mix the liquids through a high-frequency solenoid valve at a flow rate of 1.8 mL / min for 120 s.

[0105] S18: Use a high-pressure liquid delivery pump to deliver eluent F to the separation chromatographic column, and continuously switch and mix the liquids through a high-frequency solenoid valve at a flow rate of 1.8 mL / min for 100 s.

[0106] S19: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 50 s.

[0107] As can be seen from the above examples, the present invention's eluents eliminate the need for replacement eluents when separating and measuring different hemoglobin types. Simply switching the fluid circuit timing control allows for free combination, mixing, and delivery of eluents, achieving the same effect as replacing reagents. This system not only meets the rapid testing needs for the separation and measurement of HbA1a, HbA1b, HbF, L-A1c, HbA1c, and HbA0 hemoglobin, but also meets the separation and measurement needs of HbA2, HbE, HbD, HbS, and HbC.

[0108] The present invention has multimodal applicability: by providing a variety of eluents and measurement modes, the present invention can adapt to different types of hemoglobin measurement needs, including conventional hemoglobin measurement, mutant hemoglobin measurement and thalassemia-related hemoglobin measurement.

[0109] The method of the present invention simplifies the operation process, does not need to replace the chromatographic column and eluent, and can achieve efficient separation and determination of different hemoglobins by simply changing the combination and ratio of the eluent.

[0110] The present invention is highly time-efficient: by eliminating the steps of replacing the chromatographic column and eluent, the method of the present invention greatly shortens the measurement time and improves work efficiency, making it particularly suitable for emergency and batch testing needs; the simplified operating process and reduced reagent consumption reduce the overall operating cost, making the method of the present invention more economical and affordable.

[0111] The eluent composition of the present invention is carefully designed to ensure the accuracy and repeatability of the measurement results and improve the reliability of the detection.

[0112] The eluent composition of the present invention is suitable for the determination of various hemoglobins, including but not limited to HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0, HbA2, HbE, HbD, HbS and HbC, making it a powerful tool for hemoglobin research and clinical diagnosis.

[0113] The above description is only a preferred embodiment of the present invention and does not limit the patent scope of the present invention. All equivalent structural transformations made by using the description and content of the present invention under the inventive concept of the present invention, or direct / indirect application in other related technical fields are included in the patent protection scope of the present invention.

Claims

1. A multimodal method for determining hemoglobin eluate, characterized in that: The steps include: Step X1: Select the corresponding eluent, eluent combination, and measurement mode according to the type of hemoglobin to be measured; Step X2: injecting a filler into a high-pressure liquid delivery pump and delivering the selected eluent or eluent combination into the chromatographic column using the high-pressure liquid delivery pump; Step X3: separating and measuring hemoglobin in a chromatographic column; The eluents include eluent A, eluent B, eluent C, eluent E, eluent F and eluent G for measuring hemoglobin; The raw materials of the eluent A include 0.08wt%-0.12wt% of citric acid monohydrate, 0.40wt%-0.50wt% of tri-alkali metal citrate hydrate, 0.3wt%-0.5wt% of alkali metal perchlorate and 0.01wt%-0.04wt% of sodium hydroxide; The raw materials of the eluent B include 0.01wt%-0.08wt% of citric acid monohydrate, 0.50wt%-0.60wt% of tri-alkali metal citrate hydrate, 1.0wt%-2.0wt% of alkali metal perchlorate and 0.02wt%-0.08wt% of sodium hydroxide; The raw materials of the eluent C include 0.001wt%-0.01wt% of citric acid monohydrate, 0.50wt%-0.60wt% of tri-alkali metal citrate hydrate, 0.05wt%-0.15wt% of alkali metal perchlorate and 0.02wt%-0.08wt% of sodium hydroxide; The eluent E is a mixture of eluent A and eluent C, with a weight ratio of 0.4-1.0; The eluent F is a mixture of eluent B and eluent C, with a weight ratio of 0.1 to 0.8; The eluent G is a mixture of eluent C and eluent A, with a weight ratio of 0.25 to 0.5; The assay modes include rapid mode, rapid variation mode, variation assay mode and thalassemia mode; Among them, the rapid mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c and HbA0 hemoglobin; The rapid mutation mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0 hemoglobins, and mutant hemoglobins V-Win, where mutant hemoglobins V-Win are a general term for HbE, HbD, HbS, and HbC; The variation pattern is used to separate the determination of HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0 hemoglobin and HbE, HbD, HbS and HbC; The thalassemia mode is used to separate and measure HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0, HbA2, HbE, HbD, HbS and HbC.

2. The multimodal hemoglobin eluate determination method according to claim 1, characterized in that: The filler is a methacrylate copolymer, which is used to connect a high-pressure liquid delivery pump and an eluent and to separate and measure hemoglobin; the cation exchange group of the chromatographic column is one or more of a sulfonic acid group, a carboxyl group and a phosphate group.

3. The multimodal hemoglobin eluate determination method according to claim 1, characterized in that: The specific steps in quick mode are: S1: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 2.0 mL / min and a delivery time of 25 s; S2: Use a high-pressure liquid delivery pump to deliver eluent B to the separation column at a flow rate of 2.0 mL / min and a delivery time of 10 s; S3: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 20 s.

4. The multimodal hemoglobin eluate determination method according to claim 1, characterized in that: The specific steps in the rapid mutation mode are: S4: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 30 s; S5: Use a high-pressure liquid delivery pump to deliver eluent C to the separation column at a flow rate of 1.8 mL / min and a delivery time of 25 s. S6: Use a high-pressure liquid delivery pump to deliver eluent B to the separation column at a flow rate of 1.8 mL / min and a delivery time of 15 s. S7: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 30 s.

5. The multimodal hemoglobin eluate determination method according to claim 1, characterized in that: The specific steps in the mutation mode are: S8: Use a high-pressure liquid delivery pump to deliver eluent A to the separation column: flow rate is 1.8 mL / min, delivery time is 35 s; S9: Use a high-pressure liquid delivery pump to deliver eluent C to the separation column at a flow rate of 1.8 mL / min and a delivery time of 45 s. S10: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 10 s; S11: Use a high-pressure liquid delivery pump to deliver eluent C to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 10 s. S12: Use a high-pressure liquid delivery pump to deliver eluent B to the separation column at a flow rate of 1.8 mL / min and a delivery time of 10 s. S13: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 30 seconds.

6. The multimodal hemoglobin eluate determination method according to claim 1, characterized in that: Specific steps in the thalassemia model: S14: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 35 s; S15: Use a high-pressure liquid delivery pump to deliver eluent C to the separation column at a flow rate of 1.8 mL / min and a delivery time of 25 s. S16: Use a high-pressure liquid delivery pump to deliver eluent A to the separation chromatographic column at a flow rate of 1.8 mL / min and a delivery time of 10 s; S17: Use a high-pressure liquid delivery pump to deliver eluent E to the separation chromatographic column, and continuously switch and mix the liquids through a high-frequency solenoid valve at a flow rate of 1.8 mL / min for 120 s. S18: Use a high-pressure liquid delivery pump to deliver eluent F to the separation chromatographic column, and continuously switch and mix the liquids through a high-frequency solenoid valve at a flow rate of 1.8 mL / min for 100 s. S19: Use a high-pressure liquid delivery pump to deliver the eluent G to the separation chromatographic column, and use a high-frequency solenoid valve to continuously switch and mix the liquids at a flow rate of 1.8 mL / min for 50 s.

7. The multimodal hemoglobin eluate determination method according to claim 1, characterized in that: The eluent A has an osmotic pressure of 100 mOsm-200 mOsm, a pH of 5.0-6.0, and a conductivity of 8.2 sm / cm-8.6 sm / cm; The eluent B has an osmotic pressure of 500 mOsm-700 mOsm, a pH of 6.0-7.0, and a conductivity of 25 sm / cm-35 sm / cm; The eluent C has an osmotic pressure of 10 mOsm-200 mOsm, a pH of 6.0-7.0, and a conductivity of 3.0 sm / cm to 6.0 sm / cm.

8. The multimodal hemoglobin eluate determination method according to claim 1, characterized in that: The hemoglobin is selected from one or more of HbA1a, HbA1b, HbF, L-A1c, HbA1c, HbA0, HbA2, HbE, HbD, HbS and HbC.