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Fipronil artificial hapten, synthetic method for the same, and its antigen, antibody and use

A technology of artificial haptens and artificial antigens, which is applied in the direction of animal/human proteins, material inspection products, albumin peptides, etc., can solve the problems of slow metabolism, high toxicity and side effects, and high toxicity, and achieve high detection sensitivity and good affinity

Inactive Publication Date: 2008-01-09
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In particular, the pesticide is highly toxic (very toxic) to aquatic crustaceans fish, shrimp, and crabs, and also quite toxic (highly toxic) to bees, and metabolizes slowly in the environment. Two metabolites are toxic It is more than 10 times more toxic to mammals than the parent pesticide, and has an enrichment effect in the fat of the organism. It is a pesticide with relatively toxic and side effects in the ecological environment, and it needs to be strictly monitored.

Method used

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  • Fipronil artificial hapten, synthetic method for the same, and its antigen, antibody and use
  • Fipronil artificial hapten, synthetic method for the same, and its antigen, antibody and use
  • Fipronil artificial hapten, synthetic method for the same, and its antigen, antibody and use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1 Synthesis of fipronil hapten:

[0026] A kind of fipronil artificial hapten FHS, the molecular structure is:

[0027]

[0028] It is used as a raw material for the antigen system of animal immunity.

[0029] The synthetic method of the artificial hapten FHS of fipronil is as follows:

[0030] Add 4.51g 96.7% fipronil technical drug (0.01mol), 3.0g succinic anhydride (0.03mol), 200ml pyridine (dry and re-vaporized with KOH) into a 250ml three-necked flask, stir well and add 0.6g4- Dimethylaminopyridine (DMAP), the reaction system is evacuated, and N is introduced 2 It is heated and magnetically stirred at 60-70°C for 24 hours. After the reaction solution was cooled, the solvent was evaporated by rotary evaporation to obtain a brown oil. 50ml of dilute hydrochloric acid was added to remove the remaining pyridine. Then 30ml of ethyl acetate was added to dissolve the oil, and then transferred to a 250ml separatory funnel for layering. Extract twice with ethyl acetat...

Embodiment 2

[0037] Example 2 Preparation of artificial antigen and antibody of fipronil:

[0038] An artificial antigen of fipronil, the molecular structure is

[0039]

[0040] This artificial fipronil antigen is prepared by coupling the fipronil hapten FHS and protein. The molecular structure of the hapten FHS is: The binding ratio of the hapten compound to the protein is 5:1-100:1, and the protein can be bovine serum albumin or ovalbumin.

[0041] A fipronil specific antibody is a monoclonal or polyclonal immunoglobulin G that is obtained by immunizing white mice or rabbits with the artificial antigen of fipronil and can react specifically with fipronil. Used to detect fipronil residues in food, agricultural products and environmental samples; environmental samples are soil samples or water samples.

[0042] The method for preparing the artificial antigen and antibody of fipronil is as follows:

[0043] 1. Synthesis and purification of immunogen

[0044] The synthesis of immunogen utili...

Embodiment 3

[0069] Example 3 Establishment and identification of fipronil enzyme-linked immunosorbent assay method:

[0070] 1. Establishment of fipronil ELISA method and its working conditions and basic parameters

[0071]The hapten FHS and antibody are respectively covalently coupled with horseradish peroxidase to prepare enzyme-labeled hapten and enzyme-labeled antibody. Coat the polystyrene microplate with the coated antigen or antibody, add the mixture of the sample to be tested (or fipronil standard sample) and the enzyme marker, fipronil, the enzyme marker and the coating on the microtiter The coating source or antibody on the surface of the well undergoes a competitive immune binding reaction, washing to remove free substances, adding enzyme substrate and color reagent, the strength of the enzymatic color reaction and the enzyme label bound to the coating source or antibody The amount is directly proportional to the content of fipronil in the sample (or standard sample). Therefore, th...

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Abstract

A flu-insect nitrile artificial semi-antigen, its synthesis and specific antibody made from it are disclosed. The molecular structural formula of semi-antigen is (I); the molecular structural formula of specific antibody is (II). It can be used to inspect flu-insect residues in sample.

Description

Technical field [0001] The invention relates to the use of selecting a compound with -COOH and most likely to contain the original structure of fipronil as an artificial antigen, artificial antibody and antibody prepared by fipronil hapten. Background technique [0002] The invention belongs to the technical field of immunochemistry and residue analysis of pesticide small molecule compounds (molecular weight less than 1000 Daltons), involving organic synthesis, immunochemistry and biochemistry, etc., relying on the basic principles of immunology, immunochemistry and biotechnological means to design and synthesize The small molecule target analyte hapten is coupled with the carrier protein to prepare effective artificial antigens. The immunized animal prepares specific antibodies to the small molecule analyte, using the specific immunological reaction of the antigen antibody and the identification of the markers that can be easily detected. Amplification, quantitative detection of...

Claims

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Application Information

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IPC IPC(8): C07D231/44C07K14/76G01N33/53
Inventor 郭逸蓉程敬丽王春梅朱国念
Owner ZHEJIANG UNIV
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