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Kit for using proximity ligation assay to quantitatively assay Cys C content, preparation method and usage

A connection technology and quantitative detection technology, applied in the field of ortho-ligation technology, can solve the problems of poor accuracy of detection results, poor stability of latex reagents, and low detection sensitivity of enzyme-linked immunosorbent assay method.

Inactive Publication Date: 2018-06-29
NANJING TZONE BIOLOGICAL SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods have corresponding shortcomings. The detection sensitivity of enzyme-linked immunosorbent assay is low, and the stability of liquid latex reagent in immunoturbidimetric method is not good, and the accuracy of detection results is not good.

Method used

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  • Kit for using proximity ligation assay to quantitatively assay Cys C content, preparation method and usage
  • Kit for using proximity ligation assay to quantitatively assay Cys C content, preparation method and usage
  • Kit for using proximity ligation assay to quantitatively assay Cys C content, preparation method and usage

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Preparation of Cys C probe A:

[0026] 1) Take 0.1mg of anti-Cys C antibody and incubate with DBCO-sulfo-NHS reagent for 5-30min;

[0027] 2) After the reaction is over, add 10 μL of 0.5mol / L Tris-HCl and incubate for 5-10 minutes to terminate the reaction;

[0028] 3) Add the above-mentioned modified antibody to oligodeoxynucleotide a, and incubate overnight at 4°C to obtain Cys C probe A.

Embodiment 2

[0030] Preparation of Cys C probe B:

[0031] 1) Take 0.1mg of anti-Cys C antibody and incubate with DBCO-sulfo-NHS reagent for 5-30min;

[0032] 2) After the reaction is over, add 10 μL of 0.5mol / L Tris-HCl and incubate for 5-10 minutes to terminate the reaction;

[0033] 3) Add the above-mentioned modified antibody to oligodeoxynucleotide b, and incubate overnight at 4°C to obtain Cys C probe B.

Embodiment 3

[0035] The main components of the kit:

[0036] 1) Cys C probe A;

[0037] 2) Cys C probe B;

[0038] 3) connection buffer;

[0039] 4) qPCR reaction solution.

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Abstract

The invention provides a kit for using proximity ligation assay to quantitatively assay Cys C content. The kit comprises a Cys C probe A, a Cys C probe B, a ligation buffer, and a qPCR reaction solution. The invention further discloses a preparation method for the kit for using proximity ligation assay to quantitatively assay Cys C content, and the method comprises: preparation of the Cys C probeA and preparation of the Cys C probe B. Finally, the invention further discloses the usage of the kit. According to the invention, the proximity ligation assay method is applied in the quantitative assay of Cys C for the first time; and compared with methods in the prior art, the kit provided by invention has the advantages of high sensitivity, high specificity, high throughput and the like, can increase the accuracy of early renal function impairment evaluation, and has great market value.

Description

technical field [0001] The invention relates to an adjacent junction technique used for in vitro immunodiagnosis to detect the content of Cys C in a human body, belonging to the field of disease diagnosis and detection. Background technique [0002] Cystatin C (Cys C) is a cysteine ​​protease inhibitor, also known as γ-trace protein and γ-posterior globulin, widely present in nucleated cells and body fluids of various tissues, and is a A low molecular weight, basic non-glycosylated protein with a molecular weight of 13.3KD and composed of 122 amino acid residues, which can be produced by all nucleated cells in the body with a constant production rate. Circulating Cys C is cleared only by glomerular filtration, is an endogenous marker reflecting changes in glomerular filtration rate, and is reabsorbed in the proximal convoluted tubule, but is completely metabolized after reabsorption , does not return to the blood, therefore, its blood concentration is determined by glomerul...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6804
CPCC12Q1/6804C12Q2531/113C12Q2561/101
Inventor 徐林
Owner NANJING TZONE BIOLOGICAL SCI & TECH