Method of biomarker for identifying sjogren syndrome and detection kit for identifying sjogren syndrome

A technology of Sjogren's syndrome and biomarkers, which is applied in the field of medicine, can solve the problems of being unsuitable for routine purposes, large volume of serum samples, and expensive instruments, and achieve the effects of simplified operation, short analysis time, and reasonable instrument prices

Inactive Publication Date: 2019-03-01
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 ar

Method used

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  • Method of biomarker for identifying sjogren syndrome and detection kit for identifying sjogren syndrome
  • Method of biomarker for identifying sjogren syndrome and detection kit for identifying sjogren syndrome
  • Method of biomarker for identifying sjogren syndrome and detection kit for identifying sjogren syndrome

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 1.5mL EP tube, add 40 μL of 0.3mol / 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 of chloro...

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 1.5mL EP tube, add 40 μL of 0.3mol / 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, take 80μL su...

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 1.5mL EP tube, add 40 μL of 0.3mol / 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 sample at 130...

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Abstract

The invention provides a method of a biomarker for identifying sjogren syndrome and a detection kit for identifying the sjogren syndrome. The biomarker comprises free mannose and glucose which are obtained by high performance liquid chromatography derived from pre-column 1-phenyl-5-methyl pyrazolone (PMP) in serum. The detection method is a pre-column PMP derived high performance liquid chromatography method. According to the technical scheme, the method and the detection kit have the advantages that pretreatment is easy, the analysis time is short, the instrument price is reasonable, the method is compliance with conventional use, operation steps are simple and easyto learn, the accuracy of detection results is high, normal people can be distinguished from sjogren syndrome patients onlyby collecting, the amount of required serum is small, and the amount of the collected blood is less than 1 ml. The results show that the analytical method can rapidly quantify the free mannose and glucose in the serum of the sjogren syndrome patients, the method and the detection kit are of great significance for studying of the relationship between serum free mannose and glucose and sjogren syndrome, and seeking for new sjogren syndrome 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 biomarkers of Sjögren's syndrome and a detection kit thereof. Background technique [0002] Sjogren's syndrome is a diffuse connective tissue disease characterized by invasion of lacrimal glands, salivary glands and other exocrine glands, with lymphocyte infiltration and specific autoantibodies. [0003] The main clinically relevant auxiliary examinations for Sjögren's syndrome are blood and urine routine examinations and other routine examinations, autoantibody examinations, lacrimal gland function examinations, salivary gland function examinations, lip gland biopsy, etc. One of the most diagnostic significance is the autoantibody test, that is, anti-SSA and anti-SSB tests. The positive rates of anti-SSA and anti-SSB antibodies are 70% and 40%, respectively, both of which have a certain false positive rate, and the former has high sensitivi...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06G01N2030/067
Inventor 张丽娟王喆贾绍友
Owner THE AFFILIATED HOSPITAL OF QINGDAO UNIV
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