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Method for identifying bio-marker of coronary heart disease and detection kit of method

A technology of biomarkers and kits, applied in the field of medicine, can solve the problems of unsuitability for routine purposes, expensive instruments, large volume of serum samples, etc., and achieve the effects of simplified operation, reasonable price of instruments, and short analysis time.

Inactive Publication Date: 2019-03-19
THE AFFILIATED HOSPITAL OF QINGDAO UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, when using enzymatic method, gas-liquid chromatography and high-resolution liquid chromatography, in order to avoid the influence of high-concentration glucose, it needs to be removed, resulting in cumbersome pretreatment process; gas-liquid chromatography-mass spectrometry instruments are expensive and not Suitable for routine purposes; large serum sample volume required

Method used

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  • Method for identifying bio-marker of coronary heart disease and detection kit of method
  • Method for identifying bio-marker of coronary heart disease and detection kit of method
  • Method for identifying bio-marker of coronary heart disease and detection kit of method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] (1) Precisely weigh mannose (Man), glucosamine (GlcN), galactosamine (GalN), glucuronic acid (GlcUA), glucose (Glc), galactose (Gal), xylose (Xyl), rock Add appropriate amount of algalose (Fuc), add deionized water to prepare two mixed standard solutions containing the above monosaccharide 0.1mg / mL, and prepare immediately for use;

[0054] (2) Create an alkaline environment: Take 40 μL of the mixed monosaccharide standard in a 1.5mL EP tube, add 40 μL of 0.3mol / L sodium hydroxide, and vortex to mix;

[0055](3) PMP derivatization: Add 60 μL 0.5mol / L 1-phenyl-5-methylpyrazolone (PMP) to each sample, vortex and mix well, and react in an oven at 70°C for 1 h after centrifugation;

[0056] (4) Neutralization reaction by adding acid: Take out the samples in the oven, let them cool down, add 40 μL of 0.3mol / L hydrochloric acid to each sample, and vortex to mix;

[0057] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chlorof...

Embodiment 2

[0081] (1) Accurately weigh the appropriate amount of mannose (Man), rhamnose (Rha) and glucose (Glc), add deionized water to prepare 5 parts of the same mixed standard solution containing the above monosaccharide 0.1mg / mL, and use it immediately match;

[0082] (2) Create an alkaline environment: Take 40 μL of the mixed monosaccharide standard in a 1.5mL EP tube, add 40 μL of 0.3mol / L sodium hydroxide, and vortex to mix;

[0083] (3) PMP derivatization: Add 60 μL 0.5mol / L PMP to each sample, vortex and mix well, and react in an oven at 70°C for 1 h after centrifugation;

[0084] (4) Neutralization reaction by adding acid: Take out the samples in the oven, let them cool down, add 40 μL of 0.3mol / L hydrochloric acid to each sample, and vortex to mix;

[0085] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chloroform, keep the supernatant, repeat 3 times;

[0086] (6) Centrifuge the sample at 13000r / min for 10min, take 80μL su...

Embodiment 3

[0108] (1) Accurately weigh the appropriate amount of mannose and glucose, add deionized water to prepare the above monosaccharides containing 0.5mg / mL, 0.25mg / mL, 0.1mg / mL, 0.05mg / mL, 0.01mg / mL, 0.005mg / mL, 0.0025mg / mL, 0.001mg / mL, 0.0005mg / mL mixed standard solution, ready-to-use;

[0109] (2) Create an alkaline environment: Take 40 μL of the mixed monosaccharide standard in a 1.5mL EP tube, add 40 μL of 0.3mol / L sodium hydroxide, and vortex to mix;

[0110] (3) PMP derivatization: Add 60 μL 0.5mol / L PMP to each sample, vortex and mix well, and react in an oven at 70°C for 1 h after centrifugation;

[0111] (4) Neutralization reaction by adding acid: Take out the samples in the oven, let them cool down, add 40 μL of 0.3mol / L hydrochloric acid to each sample, and vortex to mix;

[0112] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chloroform, keep the supernatant, repeat 3 times;

[0113] (6) Centrifuge the sample at 130...

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Abstract

The invention provides a method for identifying a bio-marker of coronary atherosclerotic heart disease (or coronary heart disease, CAD) and a detection kit of the method. The bio-marker is free mannose and glucose obtained by high performance liquid chromatography derived from pre-column PMP in serum. The detection method is a high performance liquid chromatography method derived from pre-column 1-phenyl-5-methyl pyrazolone (PMP). The technical scheme has the advantages of simple pretreatment, short analysis time, reasonable instrument price, compliance with conventional use, simple and easy-to-learn operation steps, high accuracy of detection results, capability of distinguishing normal people from CAD patients by only taking blood, minimal amount of the serum required, less than 1 mL ofthe blood taken and the like. The results show that this analysis method can rapidly quantify the free mannose and the glucose in the serum of the CAD patients, so that the method is of great significance for studying the relationship between the free mannose and the glucose in the serum and CAD and finding new CAD clinical detection markers.

Description

technical field [0001] The invention belongs to the technical field of medicine, and in particular relates to a method for identifying a CAD biomarker and a detection kit thereof. Background technique [0002] Coronary heart disease has become the biggest threat to the health of people worldwide, and it is also one of the main causes of death. In recent years, although the mortality rate caused by coronary heart disease has declined in developed countries, in developing countries such as China, the incidence and mortality of coronary heart disease are still rising steadily, seriously endangering the health of Chinese people. Most patients with early coronary heart disease have no obvious symptoms. Once onset, they may develop into acute diseases such as myocardial infarction and sudden death. Therefore, reasonable early screening is the key to improving the cure rate and quality of life of patients with coronary heart disease. At present, the diagnosis of coronary heart dis...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06G01N2030/067
Inventor 张丽娟徐颖婕于玲范
Owner THE AFFILIATED HOSPITAL OF QINGDAO UNIV
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