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Enzyme linked immunosorbent assay kit and preparation method thereof

A detection kit and enzyme-linked technology, applied in the field of biomedicine

Inactive Publication Date: 2012-01-18
北京健平九星生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Houck et al. (J Biol. Chem. 1992, 267:26031) reported an ELISA method for detecting VEGF based on colorimetry, but its detection sensitivity was only ng / ml

Method used

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  • Enzyme linked immunosorbent assay kit and preparation method thereof
  • Enzyme linked immunosorbent assay kit and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Examples of the kit of the present invention may be the following substances:

[0024] (1) VEGF monoclonal antibody coated ELISA plate, and

[0025] (2) 1 positive control (VEGF protein freeze-dried powder);

[0026] (3) One negative control (freeze-dried animal serum powder);

[0027] (4) Sample diluent: 0.05% Tween-20, 1% bovine serum albumin, 10mM disodium edetate, 0.05% thimerosal, 0.4M NaCl, pH6.5, 1 bottle;

[0028] (5) Enzyme conjugate: horseradish peroxidase labeled vascular endothelial growth factor polyclonal antibody, 1 bottle;

[0029] (6) The concentrated washing solution is: 137mM NaCl, 2.7 mM KCl, 10mM Na2HPO4, 2 mM KH2PO4, 0.05% Tween-20, Ph7.4, 1 bottle;

[0030] (7) Chromogenic reagents A and B, of which A liquid level H2O2 solution, B liquid is 3,3',5,5'-tetramethylbenzidine solution, 1 bottle each;

[0031] (8) The stop solution is 2MH2SO4, 1 bottle;

[0032] (9) 2 sheets of adhesive tape for sealing plate.

[0033] The amounts of various substances mentioned in th...

Embodiment 2

[0094] Example 2: How to use the kit

[0095] 1. Take out the required slats from the airtight bag that has been equilibrated to room temperature. Please seal the other slats and put them back to 4℃.

[0096] 2. Leave one hole as a blank hole, add 100μl (or 2 drops) of sample diluent to the other holes.

[0097] 3. In addition to the blank wells, add 100μl each of negative control wells, positive control wells and serum samples to the corresponding wells, flick and mix well. Seal the reaction wells with sealing tape at 37°C for 60 minutes.

[0098] 4. Wash the plate 3 times: (1) Automatic plate washer: Drain the liquid in the well, and the washing liquid required to be injected is 350μl. The interval between injection and suction is 15-30 seconds, and the plate is washed 3 times. (2) Manual washing of the plate: Drain off the liquid in the wells, add 350μl of washing liquid to each hole, let it stand for 30 seconds, shake off the liquid, pat dry on thick absorbent paper, and wash 3 t...

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Abstract

The invention discloses an enzyme linked immunosorbent assay kit for detecting a vascular endothelial growth factor (VEGF). The kit internally comprises an enzyme-linked reaction plate coated by a VEGF monoclonal antibody, an enzyme conjugate configured by an antibody marked by horseradish peroxidase (HRP) and a protecting agent, lyophiled-powder of VEGF protein for positive control, protein lyophiled-powder as negative control, a sample diluent, a PBST (Poly (Butylene Succinate-Co-Butylene Terephthalate)) concentrated wash solution, a chromogenic substrate configured by 3,3',5,5'-tetramethylbenzidine, 2MH2SO4 as a stop buffer and plate sealing gummed paper. The kit can be used for diagnosis and prognosis of cancer, vascular diseases, diabetes mellitus, retinopathy and rheumatoid arthritis, and has the advantages of accuracy, specificity, sensitiveness, stability, convenience and the like.

Description

Technical field [0001] The invention belongs to the technical field of biomedicine, and specifically relates to an enzyme-linked detection kit for detecting human vascular endothelial cell growth factor (VEGF). [0002] The invention also relates to a preparation method of the above kit. Background technique [0003] The development and metastasis of tumors is a multi-stage, complex, and highly selective process involving the intricate relationship between tumor cells and the host. The mechanism is still unclear. In recent years, research on the relationship between new angiogenesis and tumor development, metastasis and prognosis has progressed rapidly, proving that new angiogenesis is a necessary condition for tumor generation, metastasis and metastasis growth, and vascular endothelial cell growth factor (VEGF) is the regulation of new blood vessels The most important cytokine produced. Studies have shown that vascular endothelial cell growth factor (VEGF) has an irreplaceable r...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68
Inventor 邹检平
Owner 北京健平九星生物医药科技有限公司