Liquid chromatography-tandem mass spectrometry quantitative detection method for content of full-blood-glucose hemoglobin
A quantitative detection method, hemoglobin technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of time-consuming, high cost, and complicated operation of fully glycated hemoglobin, and achieve low cost of consumables, easy operation, and shortened pretreatment the effect of time
Active Publication Date: 2021-10-01
昆明和合医学检验所有限公司
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Problems solved by technology
[0007]
In order to solve the technical problems of complex operation, long time-consuming and high cost in the determination of full glycated hemoglobin by mass spectrometry, the present invention provides a liquid chromatography tandem mass spectrometry quantitative detection method for the content of full glycated hemoglobin. Whole blood samples were prepared as hemolyzate, hemolyzed by recombinant carboxypeptidase B, and then separated and quantitatively analyzed by HPLC-MS / MS, and HbA
0
Dipeptide Standards and HbA
1c
The mixture of dipeptide standards was used as a calibrator, and HPLC-MS / MS was used for separation and quantitative analysis, and a standard curve was drawn, using HbA in the whole blood sample to be tested.
1c
Dipeptide and HbA
0
The peak area value of the dipeptide is calculated by the standard curve to obtain the HbA in the whole blood sample to be tested
1c
and HbA
0
The concentration of the dipeptide, and then use the following calculation formula to calculate the HbA in the whole blood sample to be tested
1c
content, the unit is mass percentage, and the calculation formula is: HbA in the whole blood sample to be tested
1c
content = HbA
1c
Dipeptide concentration / (HbA
1c
Dipeptide concentration+HbA
0
Dipeptide concentration) × 100%, in the formula, HbA
1c
The dipeptide concentration represents the HbA in the whole blood sample to be tested
1c
Concentration of dipeptide, HbA
0
The dipeptide concentration represents the HbA in the whole blood sample to be tested
0
dipeptide concentration
Method used
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[0065] Example The present invention provides a quantitative detection method for the content of fully glycated hemoglobin by liquid chromatography tandem mass spectrometry
[0066] This implementation case is applicable to the determination of the glycated hemoglobin content in whole blood. Before the test, it is necessary to prepare whole blood samples, testing equipment and testing reagents. The specific operation is as follows:
[0067] 1 Draw a standard curve
[0068] 1.1 Preparation of standard song stock solution:
[0069] 1.1.1 Preparation of standard stock solution:
[0070] Accurately weigh 3mg HbA respectively 0 Dipeptide standard and 3mg HbA 1c Dipeptide standards were dissolved in 3mL of pure water, and prepared into HbA with a concentration of 1000μg / mL in pure water. 0 Dipeptide standard stock solution and HbA at a concentration of 1000 μg / mL 1c Dipeptide standard stock solution (converted according to the purity of the standard to the concentration of th...
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The invention relates to a liquid chromatography-tandem mass spectrometry quantitative detection method for the content of full-glycosylated hemoglobin, which comprises the following steps: carrying out centrifugal separation on a whole blood sample to obtain blood cells, preparing the blood cells into hemolytic fluid, breaking hemoglobin at a second amino acid residue (His) at the N tail end of a [beta] chain under the hydrolysis action of recombinant carboxypeptidase B to obtain glycosylated HbA1c dipeptide and non-glycosylated HbA0 dipeptide, and carrying out liquid chromatography-tandem mass spectrometry on the non-glycosylated HbA0 dipeptide to obtain the content of full-glycosylated hemoglobin. The dipeptides are separated and quantified by HPLC-MS / MS (High Performance Liquid Chromatography-Mass Spectrometry / Mass Spectrometry), a mixture of an HbA1c dipeptide standard substance and an HbA0 dipeptide standard substance is used as a calibrator for separation and quantification to obtain a standard curve, and the contents of the HbA1c dipeptide and the HbA0 dipeptide are calculated according to the peak areas of the HbA1c dipeptide and the HbA0 dipeptide. The detection method has the advantages of small required sample size, high sensitivity, strong specificity, good reproducibility and short analysis time, and can provide reliable basis for clinical diagnosis.
Description
technical field [0001] The invention relates to the field of medical testing, in particular to a liquid chromatography tandem mass spectrometry quantitative detection method for the content of fully glycated hemoglobin. Background technique [0002] Glycosylated hemoglobin (Hemoglobin A 1c ; HbA 1c ) is a stable compound in which glucose in human blood is covalently bonded to the N-terminal valine residue of the hemoglobin β chain, and its full name is: hemoglobin β chain (blood)-N-(1-deoxyfructo-1-yl) hemoglobin beta chain. HbA 1C The free aldehyde group of glucose and the amino group of the N-terminal valine of the β-chain of HbA undergo a non-enzymatic combination reaction to form an unstable Schiff base (aldimine), then undergo Amadori (glucosamine) rearrangement, and finally A stable ketamine compound is formed, and its content mainly depends on the blood sugar concentration and the contact time between blood sugar and hemoglobin, which can reflect the average blood...
Claims
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Patent Timeline
Login to View More IPC IPC(8): G01N30/02G01N30/06G01N30/72
CPCG01N30/02G01N30/06G01N30/72
Inventor 吕小波夏跃莲黄和飞李佳兴谷叶
Owner 昆明和合医学检验所有限公司