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Enzyme-linked immunosorbent assay kit for detecting residual zilpaterol and use method thereof

A technology of enzyme-linked immunosorbent reagent and enzyme-linked plate, which is applied in the field of enzyme-linked immunosorbent assay and enzyme-linked immunosorbent assay kit, can solve the problems of complex operation, difficult promotion, expensive equipment, etc.

Inactive Publication Date: 2014-05-21
JIANGSU WISE SCI & TECH DEV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] In the prior art, detection methods for zilpaterol are mainly high-performance liquid chromatography-tandem mass spectrometry, high-performance liquid chromatography, etc., but the detection process of these methods is cumbersome, the detection time is long, the equipment required is expensive, and the operation is complicated. Difficult to promote

Method used

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  • Enzyme-linked immunosorbent assay kit for detecting residual zilpaterol and use method thereof
  • Enzyme-linked immunosorbent assay kit for detecting residual zilpaterol and use method thereof

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Embodiment approach

[0047] According to a preferred embodiment of the present invention, the operating steps of the inventive method are:

[0048] Take the 96-well plate coated with zilpaterol antigen, add zilpaterol standard and processed sample 50 μL / well to the corresponding microwell; add zilpaterol antibody working solution, 50 μL / well Gently oscillate to mix well, cover the plate with the cover film and place it in a light-proof environment at 25°C for 30 minutes; carefully uncover the cover film, dry the liquid in the well, and wash thoroughly with 250 μL / well of washing working solution 4~5 times, each interval of 10 s, pat dry with absorbent paper; add zilpaterol enzyme-labeled secondary antibody 100 μL / well, shake gently to mix, cover the plate with a cover film, and place it in a dark environment at 25°C React in medium for 30 min, take out and wash the plate repeatedly as above; add 100 μL / well of substrate solution, shake and mix gently, cover the plate with a cover film and place it...

Embodiment 1

[0065] Example 1 Preparation of Zipaterol-ELISA Kit

[0066] Zipaterol is a small molecule hapten, which has only reactogenicity but no immunogenicity. It needs to be combined with macromolecular substances before it can be used to immunize animals to prepare antibodies.

[0067] Zipaterol drug is a small molecular substance, which is a hapten. It only has reactogenicity, but no immunogenicity. It needs to be linked with a macromolecular carrier protein to be immunogenic. Take zilpaterol, modify the amino active group in its molecular structure, and couple with carrier protein to prepare the public antigen against zilpaterol. In this study, zilpaterol was used as hapten and carrier protein (BSA, etc.) to prepare artificial antigen by carbodiimide method.

[0068] Preparation of zilpaterol-BSA antigen:

[0069] Mix zilpaterol hapten and carrier protein BSA in 0.05 mol / L pH 9.6 carbonate buffer solution (CBS) according to the binding ratio of 11:1, then add carbodiimide, stir ...

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Abstract

The invention discloses an enzyme-linked immunosorbent assay kit for detecting residual zilpaterol and a use method thereof, and belongs to the technical field of enzyme-linked immunosorbent assay (ELISA). The kit includes an ELISA plate coated with a zilpaterol antigen, a zilpaterol standard substance working solution, a zilpaterol antibody working solution, a zilpaterol enzyme-labeled second antibody working solution, a substrate solution, a stopping liquid, a concentrated washing liquid and a concentrated sample diluent. The kit adopts an indirect competitive ELISA, and residual zilpaterol in a standard substance or a to-be-detected sample and a pre-coated antigen on the ELISA plate jointly compete for the zilpaterol antibody. The method can be used for direct detection of zilpaterol in animal derived foods, urine samples, animal tissues and serum samples, has the characteristics of convenience, rapidness, sensitivity and the like, and is suitable for detection of samples on a large scale; and the sensitivity degree of the kit is 0.1 ng / mL.

Description

technical field [0001] The invention discloses an enzyme-linked immunosorbent assay kit for detecting zilpaterol in animal tissues and its metabolites and a detection method thereof, belonging to the technical field of enzyme-linked immunosorbent assay (Enzyme-Linked Immunosorbent Assay, ELISA). The invention relates to an enzyme-linked immunoimmunoassay kit for detecting residual zilpaterol content in animal source food, blood and urine and a detection method using the kit. Background technique [0002] beta 2 - Adrenergic receptor agonists can increase the muscle / fat ratio, and when such substances are used in animal breeding, they can significantly increase the lean meat rate of reared animals. However, if it is used in large quantities or in excess for a long time, and the safe withdrawal period is not observed, it will accumulate in animal tissues, and human consumption of this animal tissue will cause poisoning or even death. Therefore, many countries and internation...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N33/531G01N21/31G01N1/28
CPCG01N33/9433G01N33/543
Inventor 杜道林洪霞
Owner JIANGSU WISE SCI & TECH DEV
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