Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

DiHT-type universal hapten and artificial antigen of food radiation marker, and preparation method and application thereof

A hapten and antigen technology, which is applied in the field of food irradiation marker DiHT general hapten, artificial antigen and its preparation, to achieve the effect of less side reactions and easy operation

Inactive Publication Date: 2012-09-12
INST OF AGRO FOOD SCI & TECH CHINESE ACADEMY OF AGRI SCI +1
View PDF4 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are few reports on the preparation of synthetic antibodies and artificial antigens using dihydrodeoxythymidine (DiHT) derivatives as haptens for enzyme-linked immunoassay (ELISA)

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • DiHT-type universal hapten and artificial antigen of food radiation marker, and preparation method and application thereof
  • DiHT-type universal hapten and artificial antigen of food radiation marker, and preparation method and application thereof
  • DiHT-type universal hapten and artificial antigen of food radiation marker, and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] Example 1. Hapten preparation and identification

[0058] Weigh 5 mg of DiHT and 16.4 mg of succinic anhydride, and dissolve them in 5 mL of anhydrous pyridine. Heated at 60°C, stirred, and refluxed for 10h. After the reaction, add 10 mL of dichloromethane to the solution, collect the lower layer mixture, then wash 3 times with 10 mL of 5% (v / v) hydrochloric acid, discard the dichloromethane layer, dissolve the residue with 10 mL of ether, and wash with 10 mL of deionized Washed with water three times, the ether layer was dried over anhydrous magnesium sulfate, vacuum filtered, purified by silica gel column chromatography, and the ether was volatilized to obtain an oily substance. Adopt thin-layer chromatography (TLC) to follow up in reaction process, and to product nuclear magnetic resonance collection of books ( 1 H-NMR) for characterization, see the process figure 2 .

Embodiment 2

[0059] Synthesis of embodiment 2.DiHT artificial antigen

[0060] The hapten DiHT was coupled to carrier proteins BSA and OVA by mixed anhydride method and activated ester method, respectively.

[0061] (1) Preparation of artificial antigen DiHT-COOH-BSA method by mixing anhydrides

[0062] Weigh 6.68 mg of DiHT-COOH powder and dissolve it in 3 mL of anhydrous DMF, oscillate and stir, add 40 μL of tri-n-butylamine, add 40 μL of isobutyl chloroformate under ice-cooling, stir and react at 4°C for 1 hour, and then warm to room temperature for 1 hour. It is called reaction solution 1. Weigh 120mg of BSA dissolved in 12mL of 0.2mol / L borate buffer (pH=8.7), this is solution 2. The reaction solution 1 was added dropwise to the solution 2 under stirring at room temperature, and stirred overnight at 15°C. The mixed solution was stirred and dialyzed with distilled water at 4°C, and the water was changed once every 4 hours for a total of 6 times until the hapten UV absorption was not...

Embodiment 3

[0065] Example 3. Identification of artificial antigens

[0066] The identification of artificial antigen is mainly carried out by two methods of UV ultraviolet scanning and SDS-PAGE gel electrophoresis.

[0067] (1) UV scanning identification

[0068] Dilute the artificial antigen and BSA to an appropriate concentration, scan in the wavelength range of 190-400nm, and obtain the ultraviolet spectrum of the hapten.

[0069] (2) Identification by SDS-PAGE gel electrophoresis

[0070] Select the concentration of the stacking gel to be 5%, the concentration of the separating gel to be 12%, the voltage of the stacking gel to be 120v, and the voltage of the separating gel to be 60v. Photographic observation.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention provides a DiHT-type universal hapten and a DiHT-type artificial antigen of food radiation marker, and a preparation method and application thereof. The preparation method comprises the steps of: taking dihydrothymine (DiHT) as an initial reactant, synthesizing a derivative of dihydrothymine by using a succinic anhydride method, wherein the derivative is hapten DiHT-COOH; coupling the hapten with carrier proteins BSA, OVA by adopting a mixed anhydride method and an active ester method, preparing a dihydrothymine artificial antigen and a dihydrothymine coating antigen, preparing a polyclonal antibody serum through BALB / c mouse immunized by the artificial antigen. A titer of the polyclonal antibody serum reaches 1:2.56*10<4> assayed by an indirect uncompetitive ELISA method, and an IC50 of the polyclonal antibody serum for DiHT is 191.8669 ng / mL, thereby indicating that the obtained DiHT antibody serum has a high affinity for DiHT-type materials. According to the invention, the yield is high, the effect is good, the operation is convenient and quick, and a base for establishing a food radiation immunological identification technology based on DiHT, the specific products of irradiated food, is provided.

Description

technical field [0001] The invention belongs to the technical field of immunoassay and biochemical industry, and relates to a food irradiation marker DiHT general hapten, an artificial antigen and a preparation method and application thereof. Background technique [0002] After decades of research, irradiation technology, as an efficient, ecological, and safe high-tech, plays an irreplaceable role in ensuring food hygiene and safety, reducing post-production losses, and promoting trade and consumption. Its application has been recognized by more and more countries. According to preliminary statistics, 38 countries and regions around the world have approved 548 kinds of food and condiments that can be treated with irradiation. However, in view of the particularity of irradiated food, the general standards of CAC irradiated food and the laws and regulations of many countries stipulate that irradiated food should be labeled. The general rules of CAC irradiated food stipulate th...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): C07H19/073C07K14/765C07K14/77C07K1/113C07K16/44C07K16/06C12N5/20G01N21/33G01N24/08G01N33/561
Inventor 李淑荣王志伟潘家荣王志东高美须李秉超睢珂王磊
Owner INST OF AGRO FOOD SCI & TECH CHINESE ACADEMY OF AGRI SCI
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com