Single-chain antibody against fenitrothion and preparation method thereof

A technology for single-chain antibodies and fenitrothion, which is applied in the field of detection antibodies, can solve the problems of no preparation of anti-fenitrothion single-chain antibodies, and achieve enhanced library capacity and molecular diversity, improved efficiency, and high specificity sexual effect

Inactive Publication Date: 2011-11-02
CHONGQING UNIV
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  • Abstract
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AI Technical Summary

Problems solved by technology

[0005] At present, there is no relevant report on the preparation of anti-fenitrothion single chain antibody

Method used

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  • Single-chain antibody against fenitrothion and preparation method thereof
  • Single-chain antibody against fenitrothion and preparation method thereof
  • Single-chain antibody against fenitrothion and preparation method thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0029] (1) Preparation of fenitrothion antigen

[0030] Dissolve 5 g of fenitrothion in 40 ml of ether, pour it into a separatory funnel, and use 30 ml of cold 1% Na 2 CO 3 The solution was washed 3 times. Add 40ml of acetic acid-hydrochloric acid solution (volume ratio: 9:1) to the upper solution, and add 8g of zinc powder several times. The yellow reaction mixture was heated and stirred under reflux for 50 min; cooled, filtered, and the precipitate was washed with carbon tetrachloride, the filtrate was combined, extracted twice with carbon tetrachloride, and the filtrate was extracted with 40 ml of double distilled water. After standing for stratification, the upper layer was removed, and the lower layer was anhydrous Na 2 SO 4 Remove water. The next day, after filtration, the brown oily amino-fenitrothion was obtained. Dissolve 500 mg of amino-fenitrothion in 50 ml of distilled water containing 10 mmol of hydrochloric acid, cool, and slowly add cold 0.1 mol / L NaNO 2 ...

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Abstract

The invention relates to a single-chain antibody against fenitrothion and a preparation method thereof. A ribosome display library of a whole set of single-chain antibody genes against the fenitrothion is constructed by using a ribosome display technology and rounding a hybrid tumor technology, and three single-chain antibody genes with high affinity and high specificity for the fenitrothion are screened from the library. After the single-chain antibody genes are connected with vectors PPOW3.0 and then subjected to soluble expression in escherichia coli, three soluble proteins with molecular weights of about 30KDa are obtained; and by enzyme-linked immuno sorbent assay (ELISA) and Biacore analysis, the three antibody proteins have high affinity and high specificity for the fenitrothion, so excellent single-chain antibody genes and antibody proteins are provided for establishing ELISA and immune sensors. The invention has significance for realizing quick, simple, convenient and accurate detection of organic phosphorus pesticide residue, and provides a new direction.

Description

technical field [0001] The invention relates to an antibody for detection, in particular to an anti-fenitrothion single-chain antibody and a preparation method thereof. Background technique [0002] With the development of society and economy and the improvement of people's living standards, food safety issues have attracted widespread attention from the whole society. However, food poisoning incidents caused by excessive pesticide residues have occurred from time to time, so the detection of pesticide residues is particularly important. . Fenitrothion is one of the organophosphorus pesticides widely used at present. Over the years, methods such as gas chromatography, high performance liquid chromatography, and mass spectrometry have been successfully applied to pesticide analysis, which can accurately detect the amount of residual pesticides. However, they require expensive instruments and equipment, complicated pretreatment, skilled technicians and long analysis period; t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/44C12N15/13C12N15/10C12N15/70C12R1/19
Inventor 夏玉先罗义辉
Owner CHONGQING UNIV
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