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Preparation method and application of parathion-methyl artificial hapten, artificial antigen and specific antibody

A methyl parathion, artificial hapten technology, applied in chemical instruments and methods, specific peptides, animal/human proteins, etc., can solve the problems of expensive equipment, high personnel and site requirements, and long time.

Active Publication Date: 2012-03-14
贾明宏 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method effectively solves the defects of traditional residue detection and analysis methods such as time-consuming, expensive equipment, and high requirements for personnel and sites, and has broad application prospects

Method used

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  • Preparation method and application of parathion-methyl artificial hapten, artificial antigen and specific antibody
  • Preparation method and application of parathion-methyl artificial hapten, artificial antigen and specific antibody
  • Preparation method and application of parathion-methyl artificial hapten, artificial antigen and specific antibody

Examples

Experimental program
Comparison scheme
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preparation example Construction

[0009] Synthesis of hapten→preparation of artificial antigen→preparation of specific antibody→ELISA method establishment and

[0010] Identification → analysis of actual samples

[0011] 1. Synthesis of methyl parathion artificial hapten

[0012] Pesticide small molecules must be linked with macromolecules to stimulate animals to produce specific antibodies. Therefore, the synthesis and identification of haptens are the most basic and critical steps for the production of specific antibodies and the establishment of rapid detection technology for pesticide residues. On the one hand, the ideal hapten should have the characteristic structure of the analyte, especially the stereochemical characteristics. On the other hand, after the hapten is connected to the carrier, it should ensure that the characteristic structure of the analyte can be recognized and combined by the immune active cells to the greatest extent. To prepare antibodies with the expected selectivity. 1) Haptens ar...

Embodiment 1

[0033] Three weeks later, booster immunization was carried out, the immunization dose was 1.0 mg / kg, an equal volume of Freund's incomplete adjuvant was added, and multiple subcutaneous injections were performed on the back. After that, immunize once every three weeks with the same dose and method. From the third immunization, blood was collected from the rabbit's ear vein for one week each time, and the antibody titer and specificity were determined. Example 1: Synthesis of Hapten MP

[0034] Weigh 52.0g PSCl 3 , add 39.0g of anhydrous methanol dropwise at 15°C, react for 1h, and wash the reaction solution with water to obtain light yellow oil I→Weigh 40.0g of I, add 30.0g of p-nitrophenol, and add 3mol of tri-n-butyl under stirring Amine, after reacting for 20min, add 40mL of 6mol / L NaOH solution dropwise, and react for 1h. Filtered, extracted with ether, the ether phase was washed successively with NaOH aqueous solution with a mass fraction of 5 and distilled water, anhy...

Embodiment 2

[0037] Example 2: Synthesis of Hapten MP

[0038] Weigh 52.0g PSCl 3 , add 39.0g of anhydrous methanol dropwise at 15°C, react for 1h, and wash the reaction solution with water to obtain light yellow oil I→Weigh 40.0g of I, add 30.0g of p-nitrophenol, and add 3mol of tri-n-butyl under stirring Amine, after reacting for 20min, add 40mL of 6mol / L NaOH solution dropwise, and react for 1h. Filtered, extracted with ether, the ether phase was washed successively with NaOH aqueous solution with a mass fraction of 5 and distilled water, anhydrous MgSO 4 Drying, column chromatography, washing with petroleum ether / diethyl ether (7:1, volume ratio) to obtain light yellow oil II → weigh 6.2 g of 3-mercaptopropionic acid and 3.1 g of yellow oil II in 20 mL of dimethyl Add 2.0g of NaOH to the sulfoxide in a three-necked flask, and stir it magnetically to dissolve it; slowly heat up the oil bath to 100°C, keep it warm for 2 hours, then remove the oil bath, and after cooling to room tempera...

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Abstract

The invention discloses a preparation method and application of a parathion-methyl artificial hapten, a parathion-methyl artificial antigen and a parathion-methyl specific antibody, and belongs to the technical field of pesticide immunologic chemistry. The method comprises the following steps of: introducing spacer arms and active groups into a methoxyl site of parathion-methyl by taking 3-mercaptopropionic acid as a functional group reagent to synthesize O-methyl-N-(2-carboxyethyl) sulfo-O-(4-nitrobenzophenone) phosphorothioate (MP for short) serving as the hapten; and coupling the hapten with protein in a mixed anhydride method and an active ester method to prepare the artificial antigen. In the method, the parathion-methyl antibody with high affinity and high specificity is prepared by immunizing an animal through an immunogen, and enzyme-linked immunosorbent assay (ELISA) established by the antibody can be used for detecting trace quantity of parathion-methyl residues in a sample quickly.

Description

technical field [0001] The invention relates to a preparation method of methyl parathion artificial hapten, artificial antigen and specific antibody and its application in enzyme-linked immunosorbent assay method (ELISA). The invention belongs to the technical field of pesticide immunochemistry. Background technique [0002] The problem of environmental pollution caused by the extensive use of pesticides has aroused people's great attention. With the development of the economy and the strengthening of people's awareness of environmental protection, my country has also paid more and more attention to the monitoring of pesticide residues in the environment and food. Strengthening the research on pesticide residue monitoring has important theoretical and practical significance for the rational use of pesticides, environmental protection and human health. Traditional pesticide residue analysis methods such as liquid chromatography (HPLC), gas chromatography (GC), and mass spec...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07F9/18C07K14/765C07K14/795C07K16/06G01N33/543
Inventor 贾明宏
Owner 贾明宏
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