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Artificial antigen and antibody for immunoassay of melamine residues and preparation method thereof

A melamine, artificial antigen technology, applied in the direction of serum immunoglobulin, animal/human protein, material test, etc., can solve the problem of high price, achieve high similarity, stable antibody, avoid synthetic reaction and impurity removal process Effect

Inactive Publication Date: 2013-10-09
JIANGSU INST OF POULTRY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the melamine residue immunoassay kits and reagent strips reported in domestic literature are all imported from abroad, which are expensive

Method used

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  • Artificial antigen and antibody for immunoassay of melamine residues and preparation method thereof
  • Artificial antigen and antibody for immunoassay of melamine residues and preparation method thereof
  • Artificial antigen and antibody for immunoassay of melamine residues and preparation method thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0025] Example 1: Preparation and Identification of Melamine Artificial Antigen (Immune Antigen and Coating Antigen)

[0026] 1. Synthesis of melamine artificial antigen

[0027] Add 41.5-42.5 mg of hapten, 33.5-34.5 mg of NHS, and 61.5-62.5 mg of DCC into 2 mL of DMF, and stir overnight at 4°C. The next day, the mixture was centrifuged, the precipitate was discarded, and the supernatant was absorbed for later use. Dissolve 112.5-113.0 mg of BSA and 74.5-75.0 mg of OVA in 8 mL of 0.01 mol / L, pH=7.4 PBS, divide the obtained supernatant into two parts and slowly add the two solutions while stirring, at 4 °C Stir slowly overnight. Afterwards, the reaction solution was transferred to the treated dialysis bag, collected after thorough dialysis in PBS, subpackaged, and stored at -20°C.

[0028] 2. Identification of melamine artificial antigen

[0029] Ultraviolet spectroscopy is used for identification. After the carrier protein and the hapten are coupled, the product will prese...

Embodiment 2

[0030] Embodiment 2: Preparation and specific identification of melamine antibody

[0031] 1. Preparation of melamine antibody

[0032] (1) Immunization Three-month-old healthy, male New Zealand white rabbits with a weight of about 1.5 kg were selected and raised in a standard experimental animal room for observation for 3 days, and then immunized after confirming that their physical condition was normal. The dose of immunogen Mel-BSA is 1 mg / kg body weight, and the immunogen and adjuvant are mixed at a ratio of 1:1. For the first immunization, the immunogen is emulsified with Freund's complete adjuvant, and for the subsequent booster immunization, it is emulsified with Freund's incomplete adjuvant Immunogen, antigen and adjuvant form a "water-in-oil" state. Take a drop of antigen and drop it on the water surface. The antigen "water-in-oil" state floats on the water surface and does not disperse, indicating that the emulsification is complete. The booster immunization was car...

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Abstract

The invention discloses an artificial antigen and antibody for immunoassay of melamine residues and a preparation method thereof, belonging to the immunochemical analysis technology of small-molecular compounds. According to the invention, 4,6-diamino-1,3,5-triazine-2-acetic acid is directly used as a hapten, and the hapten is combined with bovine serum albumin or ovalbumin by an active ester method to obtain a melamine artificial antigen; and an antibody is prepared through animal immunization, wherein the antibody has good specificity to melamine, and a foundation is laid for an immunological detection method for quickly and accurately detecting melamine residues in food.

Description

technical field [0001] The invention belongs to the technical field of immunochemistry and residual analysis of small molecule compounds, and relates to the synthesis of melamine small molecule artificial antigen with aminobenzene ring molecular structure and the preparation of immune animal specific antibody. Background technique [0002] Melamine is a triazine nitrogen-containing heterocyclic organic compound. Because of its high nitrogen content, white and odorless, easy to buy and produce, inherently unscrupulous manufacturers add it to food and feed for the shortcomings of the Kjeldahl nitrogen method. Increase protein content. Although melamine is a low-toxic compound, and the half-lethal dose of oral administration to rats is greater than 3g / kg body weight, studies by the U.S. FDA have shown that melamine and calcium form insoluble complexes under the maintenance of urine pH in the body, and long-term intake of melamine by animals will cause reproductive problems. , ...

Claims

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Application Information

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IPC IPC(8): C07K14/765C07K14/77C07K16/06G01N33/68
Inventor 张小燕高玉时施祖灏陆俊贤唐梦君唐修君葛庆联蒲俊华顾荣
Owner JIANGSU INST OF POULTRY SCI
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