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Method and detection kit for identifying biomarker of cerebral infarction

A biomarker and cerebral infarction technology, applied in the field of medicine, can solve the problems that metal implants cannot be applied to patients, difficult to display calcified sites, and not suitable for routine purposes, etc., to achieve simplified operation, less serum consumption, and low instrument price reasonable effect

Inactive Publication Date: 2019-02-15
THE AFFILIATED HOSPITAL OF QINGDAO UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

MRI is a detection technique that has gradually become popular in recent years. It can sensitively reflect the changes of brain hematoma, but it also has limitations in use: 1. It is difficult to show the calcification site; 2. It is not applicable to patients with metal implants in the body
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
At present, there is no literature report on the simultaneous detection of free mannose and glucose in the serum of patients with cerebral infarction by pre-column 1-phenyl-5-methylpyrazolone (PMP)-derived HPLC

Method used

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  • Method and detection kit for identifying biomarker of cerebral infarction
  • Method and detection kit for identifying biomarker of cerebral infarction
  • Method and detection kit for identifying biomarker of cerebral infarction

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] (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;

[0053] (2) Create an alkaline environment: Take 40 μL of the mixed monosaccharide standard in a 1.5 mL centrifuge tube, add 40 μL of 0.3 mol / L sodium hydroxide, and vortex to mix;

[0054] (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;

[0055] (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;

[0056] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer o...

Embodiment 2

[0080] (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;

[0081] (2) Create an alkaline environment: Take 40 μL of the mixed monosaccharide standard in a 1.5 mL centrifuge tube, add 40 μL of 0.3 mol / L sodium hydroxide, and vortex to mix;

[0082] (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;

[0083] (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;

[0084] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chloroform, keep the supernatant, repeat 3 times;

[0085] (6) Centrifuge the sample at 13000r / min for 10min, ta...

Embodiment 3

[0106] (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;

[0107] (2) Create an alkaline environment: Take 40 μL of the mixed monosaccharide standard in a 1.5 mL centrifuge tube, add 40 μL of 0.3 mol / L sodium hydroxide, and vortex to mix;

[0108] (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;

[0109] (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;

[0110] (5) Extraction: add 500 μL chloroform to each tube, vortex, centrifuge, remove the lower layer of chloroform, keep the supernatant, repeat 3 times;

[0111] (6) Centrifuge the sam...

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Abstract

The invention provides a method and a detection kit for identifying a biomarker of cerebral infarction. The biomarker is free mannose and glucose obtained by high performance liquid chromatography based on pre-column 1-phenyl-3-methyl-5-pyrazolone (PMP) derivatization in serum. The detection method is the high performance liquid chromatography method based on the pre-column PMP derivatization. Thetechnical scheme has the advantages that the pretreatment is simple, the analysis time is short, the instrument price is reasonable, the requirements of conventional use are met, the operation stepsare simple and easy to learn, the accuracy of a detection result is high, a normal person and a cerebral infarction patient can be distinguished only by blood collection, the amount of the needed serum is extremely small, the blood collection amount is smaller than 1 mL, and the like. The obtained result shows that the analysis method can be used for rapidly determining the content of the free mannose and glucose in the serum of the cerebral infarction patient, so that the method has very important significance for researching a relation between the free mannose and glucose in the serum and the cerebral infarction, and looking for a novel cerebral infarction clinical detection marker.

Description

technical field [0001] The invention belongs to the technical field of medicine, and in particular relates to a method for identifying biomarkers of cerebral infarction and a detection kit thereof. Background technique [0002] Cerebral infarction, also known as ischemic stroke, is a cerebrovascular disease that seriously endangers the health of Chinese people. The "China Stroke Prevention Report 2016" pointed out that there are 13 million stroke patients in my country, and 1.88 million people die from stroke every year. Compared with 30 years ago, the overall mortality rate of stroke in my country has decreased, but the mortality rate of cerebral infarction has increased by 28.8%; the overall prevalence rate of stroke has risen rapidly from 0.4% in 1993 to 1.23% in 2013 Among them, the number of patients with cerebral infarction accounts for about 77.8% of the whole, and it shows a trend of younger age. Data show that about 70%-80% of patients with cerebral infarction suff...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06G01N2030/067
Inventor 张丽娟张朦张国庆裴海涛
Owner THE AFFILIATED HOSPITAL OF QINGDAO UNIV
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