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Enzyme linked immunosorbent assay kit for combined diagnosis of gastrosis or evaluation of gastric cancer risks

An enzyme-linked immunosorbent assay and kit technology, applied in anti-enzyme immunoglobulin, material separation, biochemical equipment and methods, etc., can solve the problem of high cost of human and material resources, and achieve the effect of improving sensitivity

Inactive Publication Date: 2011-06-08
BEIJING MOKOBIO LIFE SCI CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Gastric cancer screening began in Japan in the 1960s, when gastric cancer was highly prevalent. At that time, a step-by-step screening method of gastrointestinal double-contrast radiography-gastroscopy-pathological examination was adopted, which cost a lot of manpower and material resources.

Method used

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  • Enzyme linked immunosorbent assay kit for combined diagnosis of gastrosis or evaluation of gastric cancer risks
  • Enzyme linked immunosorbent assay kit for combined diagnosis of gastrosis or evaluation of gastric cancer risks
  • Enzyme linked immunosorbent assay kit for combined diagnosis of gastrosis or evaluation of gastric cancer risks

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0087] 1. Preparation of Human Pepsinogen Antigen I or Protease Antigen II Natural Protein

[0088] (1) Collect surgically resected normal human gastric tissue, rinse with phosphate buffer, separate gastric mucosa, blot dry, weigh, add phosphate buffer, mash and homogenate, centrifuge, collect supernatant, and precipitate phosphate The buffer was resuspended, centrifuged again, and the supernatant was combined to obtain the pepsinogen crude extract;

[0089] (2) Add pepsinogen crude extract to the equilibrated DEAE-52 chromatographic column, elute the chromatographic column with PBS buffer until the absorbance is 0 at 280nm, continue to elute the chromatographic column with PBS buffer , combining the elution peaks to obtain active pepsinogen;

[0090] (3) Take the peak of active protease, load it on the gel chromatography column, and use 50mmol PB-S (containing 0.05mol / LNa 2 SO 4 ) elution, the flow rate is 3.0ml / min, the pressure is 8Kpa, the elution curve shows 4 protein el...

Embodiment 2

[0099] The preparation of embodiment 2 PG I enzyme-linked immunosorbent assay kit

[0100] 1 solution preparation

[0101] a) Prepare coating buffer

[0102] Recipe: 0.05M carbonate buffer (pH=9.6)

[0103] b) Prepare blocking solution

[0104] Formulation: Phosphate buffered saline, 1% BSA, CPO 3 3- =0.01M, pH=7.4,

[0105] c) Prepare concentrated washing solution

[0106] Recipe: Phosphate buffered saline, CPO 3 3- =0.2M, pH=7.4

[0107] d) Preparation of enzyme-labeled antibody working solution

[0108] Formulation: Tris-HCL 50Mm, pH=8.5

[0109] E) Preparation of substrate A solution

[0110] Formula: urea hydrogen peroxide (0.2mg / ml), ultrapure water

[0111] F) Preparation of substrate B solution

[0112] Recipe: 0.1M citric acid-phosphate buffer (PH5.5), TMB (2mg / ml), DMSO

[0113] G) Preparation of stop solution

[0114] Recipe: 2M sulfuric acid solution

[0115] 2. Coating, sealing, vacuum packaging

[0116] 2.1 Coating

[0117] (1) Turn on the coati...

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Abstract

The invention discloses an enzyme linked immunosorbent assay kit for combined diagnosis of the gastrosis or evaluation of gastric cancer risks and a preparation method thereof. The kit comprises a micropore plate coated with an antibody against a pepsin antigen I or an antibody against a pepsin antigen II, an enzyme labeled antibody, a color-developing agent, a stop solution and a concentrated cleaning solution, wherein the pepsin antigen I or the pepsin antigen II is a natural protein obtained from extraction of human gastric mucosa tissue. The kit disclosed by the invention adopts a mouse immunized with pepsinogen I and pepsinogen II which are separated from human gastric mucosa to prepare immunogen of a monoclonal antibody, the used standard sample also adopts the pepsin antigen I or the pepsin antigen II separated from the human gastric mucosa, thereby the defects caused by adopting different structures of animal pepsinogen and human pepsinogen are filled. The kit can be used for accurately diagnosing the gastrosis or early gastric cancer and has the advantages of high sensitivity, strong specificity, good accuracy and the like.

Description

technical field [0001] The present invention relates to a kit for diagnosing diseases, in particular to an enzyme-linked enzyme-linked method for diagnosing gastric disease or early gastric cancer by measuring the concentration of pepsinogen I and pepsinogen II in a subject and comparing it with a set cut-off value. The immunoadsorption kit, and the invention also relates to a preparation method of the kit, which belongs to the field of diagnosis or detection of gastric disease or early gastric cancer. Background technique [0002] Pepsinogen (PG) is a precursor of aspartic acid protease, a single-chain polypeptide with a molecular mass of 42kDa, including three disulfide bonds, and an isoelectric point of 3.7. Human PG can be divided into 7 components according to its electrophoretic mobility, and the immunogenicity of the 1-5 components that move faster to the positive is similar, called pepsinogen I (PGI), mainly composed of the principal cells of the gastric gland and mu...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N30/06G01N33/577C12N5/20C07K16/40
Inventor 金鑫王瑞环
Owner BEIJING MOKOBIO LIFE SCI CO LTD
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