Synthesis and application of envelope antigen universal for pyrethriods pesticide

A technology of pyrethroids and antigen-coated, applied to measuring devices, instruments, scientific instruments, etc., to meet the needs of research

Inactive Publication Date: 2013-06-12
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, there is no report about the preparation of the coating agent and the ...

Method used

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  • Synthesis and application of envelope antigen universal for pyrethriods pesticide
  • Synthesis and application of envelope antigen universal for pyrethriods pesticide
  • Synthesis and application of envelope antigen universal for pyrethriods pesticide

Examples

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Embodiment 1

[0037] Example 1: Preparation of a universal coating antigen for pyrethroid pesticides

[0038] (1) Synthesis of haptens

[0039] ①Compound 3: Synthesis of N-(3-phenoxybenzoyl)-4-nitro-L-phenylalanine:

[0040] The reaction at N 2 Under protection, dissolve 500 mg of compound 1: 3-phenoxybenzoic acid in 10 mL of chloroform, place it in an ice bath after dissolving, then add 5 μL of DMF and 0.33 mL of thionyl chloride dropwise, stir for 5 min and then withdraw Remove from the ice bath, then heat in an oil bath at 65°C for 1 h, and remove chloroform and excess thionyl chloride by rotary evaporation to obtain a colorless and transparent liquid, which is dissolved by adding 1 mL of tetrahydrofuran to obtain liquid a; then add 480 mg of compound 2: Dissolve p-nitrophenylalanine and 380mg potassium carbonate in 10mL of water to form solution b; then slowly add solution a to solution b under vigorous stirring within 10 minutes, and after 1 hour of reaction, use 50mL ethyl acetate e...

Embodiment 2

[0058] 1. Preparation of coated plates

[0059] Use pH 9.6, 0.05M carbonate buffer solution to dilute the universal coating antigen of pyrethroid pesticides, doubling dilution 6 gradients, add 100 μL per well of the coating solution into a 96-well microtiter plate, overnight at 4 °C, Wash with washing solution (0.01M PBS containing 0.02% Tween20), each time pat dry on absorbent paper, and repeat four times. Then add 200 μL / well of blocking solution (pH 9.6, 0.05 M carbonate buffer solution containing 0.2% gelatin), place in a 37°C oven for 2 hours, take it out, wash it four times with the above washing solution, place in a 37°C oven for 15 minutes, and dry Store at 4°C after drying.

[0060] 2. Preparation of pyrethroid pesticide standard solution

[0061] Prepare a 1 mg / mL permethrin standard solution with n-hexane, place it in a brown glass bottle, and store it at -20°C. Before use, dilute the standard 10 times with n-hexane to 100 μg / mL. Measure 20 μL and place it in a ...

Embodiment 3

[0070]1 Coupling of gold nanoparticles and pyrethrin monoclonal antibody: The gold nanoparticles (particle size: 22-27nm) used in the present invention were synthesized by our laboratory. The specific method for coupling the group-selective antibody of pyrethroids to the surface of gold nanoparticles is: measure a certain amount of gold nanoparticle solution into a small test tube, and centrifuge (8000rpm, 10min), in order to remove citric acid in the gold nanoparticle solution Trisodium was resuspended with the same volume of ultrapure water, and then the pyrethroid group-selective antibody was slowly added dropwise into the gold nanoparticle suspension, shaken at room temperature for 2 hours, and then 1% BSA was added dropwise, so that the final concentration of BSA was 0.1%, shake at room temperature for 2 hours to block the unbound sites on the gold nanoparticles and avoid non-specific adsorption. Centrifuge (7000rpm, 8min) to remove excess antibody and BSA, resuspend in u...

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Abstract

The invention relates to synthesis and an application of an envelope antigen universal for pyrethriods pesticide, belonging to the technical field of biochemical engineering. The envelope antigen universal for pyrethriods pesticide is synthesized by taking 3-phenoxybenzoic acid and nitrobenzene alanine as raw material through two steps of chemical reaction, the envelope antigen and a carrier protein BSA (Bovine Serum Albumin) are coupled through diazotization, and the coupling ratio of a conjugate is identified and estimated through ultraviolet spectroscopy. The synthesized conjugate (envelope antigen) is used for enzyme linked immunoassay and preparation of a collaurum immunochromatography test strip. The enzyme linked immunoassay and the preparation of the collaurum immunochromatography test strip prove that the synthesis of the envelope antigen universal for pyrethriods pesticide is synthesized successfully, thus providing a basis for establishment of a immunoassay method of the pyrethriods pesticide, and meeting the domestic requirement of rapid detection on the pyrethriods pesticide.

Description

technical field [0001] The invention relates to the synthesis and application of a general-purpose coating antigen of pyrethroid pesticides. The synthesized antigen is identified by ultraviolet spectrophotometry, and is used in the preparation of enzyme-linked immunoassay and colloidal gold immunochromatography test strips , belonging to the field of biochemical technology. Background technique [0002] Pyrethroid pesticides, the English name Pyrethroid insecticides, is a class of bionic insecticides developed on the basis of the chemical structure of natural pyrethroids. It has fast insecticidal speed, short residual effect time, and low toxicity to higher animals and birds. , use safety and other advantages, are widely used in tea gardens, orchards, animal husbandry and household pest control, therefore, pyrethroid pesticides have become one of the main pillar products of modern insecticides. However, the long-term and large-scale application of the pesticide has caused s...

Claims

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Application Information

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IPC IPC(8): G01N33/531G01N33/543G01N33/577G01N33/558
Inventor 匡华陈秀金胥传来徐利广马丽萍刘丽强王利兵丁利马伟宋姗姗
Owner JIANGNAN UNIV
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