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Reagent kit for quantitatively detecting RBP content through proximity ligation assay and preparation and using methods

A connection technology and quantitative detection technology, applied in the field of adjacent connection 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

Inactive Publication Date: 2018-07-10
NANJING TZONE BIOLOGICAL SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Enzyme-linked immunosorbent assay has the characteristics of simple operation and long stable period of reagents, but the detection sensitivity of enzyme-linked immunosorbent assay is low, and immunoturbidimetry is often used in clinical practice now. This technology has become mature and the operation It is simple and has few interference factors, but the stability of the liquid latex reagent is not good, and the accuracy of the test result is not good

Method used

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  • Reagent kit for quantitatively detecting RBP content through proximity ligation assay and preparation and using methods
  • Reagent kit for quantitatively detecting RBP content through proximity ligation assay and preparation and using methods
  • Reagent kit for quantitatively detecting RBP content through proximity ligation assay and preparation and using methods

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Preparation of RBP probe A:

[0026] 1) Take 0.1mg of anti-RBP 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 modified with an azide group at the 5' end, and incubate overnight at 4°C to obtain RBP probe A.

Embodiment 2

[0030] Preparation of RBP probe B:

[0031] 1) Take 0.1mg of anti-RBP 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 modified with an azide group at the 5' end, and incubate overnight at 4°C to obtain RBP probe B.

Embodiment 3

[0035] The main components of the kit:

[0036] 1) RBP probe A;

[0037] 2) RBP probe B;

[0038] 3) connection buffer;

[0039] 4) qPCR reaction solution.

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Abstract

The invention provides a reagent kit for quantitatively detecting the RBP content through the proximity ligation assay. The reagent kit comprises an RBP probe A, an RBP probe B, a connecting buffer solution and a qPCR reaction solution. The invention further discloses a preparation method of the reagent kit for quantitatively detecting the RBP content through the proximity ligation assay. The method comprises the steps of preparing the RBP probe A and preparing the RBP probe B. The invention finally discloses a using method of the reagent kit. The using method comprises the step of firstly, applying the proximity ligation assay to RBP quantitative detection, compared with an existing technical method, the method has the advantages of being high in sensitivity and specificity, high in throughput and the like, the kidney function disease prediction accuracy can be improved, and the great market value is obtained.

Description

technical field [0001] The invention relates to an adjacent junction technique used for in vitro immunodiagnosis to detect the content of RBP in a human body, belonging to the field of disease diagnosis and detection. Background technique [0002] RetinoL Binding Protein (RBP) is a small molecular protein secreted by the liver in blood plasma. It consists of a polypeptide chain consisting of 183 amino acid residues and a small amount of carbohydrates, and does not contain neutral sugars and amino groups. Hexose, molecular weight 21,000Da. RBP is mainly synthesized by the rough endoplasmic reticulum of liver cells, widely distributed in human serum, cerebrospinal fluid, urine and other body fluids, and finally excreted in urine. It is also a transporter of vitamin A in the blood and plays an important role in the metabolism of vitamin A. Because of its short half-life, it can more sensitively reflect the disease status of the body. [0003] In recent years, RBP detection a...

Claims

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Application Information

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IPC IPC(8): G01N33/68
CPCG01N33/6893
Inventor 徐林
Owner NANJING TZONE BIOLOGICAL SCI & TECH