ELISA (Enzyme Linked Immunosorbent Assay) Kit and application thereof

An enzyme-linked immunosorbent reagent and enzyme labeling technology, applied in the field of enzyme-linked immunosorbent immunoassay kits, can solve the problems of complex instruments and equipment, unsuitable for on-site monitoring and screening of a large number of samples, and cumbersome processes

Active Publication Date: 2011-06-08
北京明日达科技发展有限责任公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The current chemical methods for detecting melamine content include high performance liquid chromatography and liquid chromatography-tandem mass spectrometry. Due to the complicated equipment and cumbersome process, they are not suitable for on-site monitoring and screening of a large number of samples.

Method used

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  • ELISA (Enzyme Linked Immunosorbent Assay) Kit and application thereof
  • ELISA (Enzyme Linked Immunosorbent Assay) Kit and application thereof
  • ELISA (Enzyme Linked Immunosorbent Assay) Kit and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Embodiment 1, preparation of melamine hapten, melamine antigen and melamine clone antibody

[0065] 1. Synthesis and identification of melamine hapten

[0066] 1. Synthesis of melamine hapten

[0067] Weigh 100mg of melamine and 100mg of maleic anhydride in a 50mL Erlenmeyer flask (the ratio of the amount of melamine to maleic anhydride is 1:1), add 5mL of dimethyl sulfoxide (DMSO), and place in a 66°C water bath In the pot, reacted for 12 hours to obtain solution I, which contained the target product. Calculate the yield of the desired product. The formula for calculating the yield is: yield=mass of product actually obtained / mass of product theoretically obtained×100%. The experiment was repeated three times, and the yield of the target product was 46.7%.

[0068] 2. Identification of melamine hapten

[0069] Get above-mentioned object product and measure its structure through ESI mass spectrometry, find that its molecular ion peak is 223 (M-1) (see figure 1 ). ...

Embodiment 2

[0101] Embodiment 2, the ELISA kit that detects melamine

[0102] 1. The enzyme-linked immunosorbent assay kit consists of the following materials:

[0103] 1. Melamine coating agent; as described in Example 1.

[0104] 2. ELISA plate;

[0105] 3. Melamine monoclonal antibody: secreted by the melamine monoclonal antibody hybridoma cell line MAL with the preservation number CGMCC No.3394;

[0106] 4. Melamine standard: the concentration of the standard solution is 0 μg / L, 5 μg / L, 20 μg / L, 100 μg / L, 500 μg / L and 2500 μg / L; the melamine standard is purchased from Sigma Aldrich; CAS number: 108- 78-1; Dilute to the above concentrations with sample diluent;

[0107] 5. Goat anti-mouse secondary antibody labeled with horseradish peroxidase;

[0108] 6. Substrate chromogenic solution: composed of liquid A and liquid B, liquid A is an aqueous solution of 2% carbamide peroxide, liquid B is an aqueous solution of 1% tetramethylbenzidine;

[0109] 7. Termination solution: 2mol / L aqu...

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Abstract

The invention discloses an ELISA (Enzyme Linked Immunosorbent Assay) kit and application thereof. The ELISA kit comprises a melamine monoclonal antibody and a melamine hapten, wherein the melamine monoclonal antibody is generated by the secretion of melamine monoclonal antibody hybridoma cell strains MAL (Myelin And Lymphocyte) with a preservation number of CGMCC (China General Microbiological Culture Collection Center) NO.3394. The ELISA kit provided by the invention is mainly used for qualitatively or quantitatively detect the content of melamine in the products (especially dairies such as milk, milk powder and the like as well as eggs, feed and the like) eaten by animals or human by the indirect competitive ELISA. By means of the ELISA Kit and the detection method, the requirement on the pretreatment of a sample is low; the pretreatment process of the sample is simple; and mass samples can be rapidly detected at the same time; by utilizing the high-specificity melamine monoclonal antibody, the detection method is convenient, is easy to carry out, and has the characteristics of high specificity, high sensitivity, high precision, high accuracy and the like.

Description

technical field [0001] The invention relates to an ELISA kit and its application. Background technique [0002] Melamine, molecular formula C 3 N 6 h 6 , referred to as triamine, is a triazine nitrogen-containing heterocyclic organic compound. Melamine is weakly alkaline and can form melamine salts with hydrochloric acid, sulfuric acid, nitric acid, acetic acid, oxalic acid, etc. Melamine is an important nitrogen-heterocyclic organic chemical raw material. In neutral or slightly alkaline conditions, it can condense with formaldehyde to form various methylol melamines, but in slightly acidic environments (pH5.5-6.5) The derivatives undergo polycondensation reaction to form resin products. Melamine is a basic organic chemical intermediate product with a wide range of uses, and its main use is as a raw material for the production of melamine-formaldehyde resin. The resin has higher hardness than urea-formaldehyde resin, non-flammable, water resistance, heat resistance, ag...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/44C12N5/20G01N33/577G01N33/04G01N33/08G01N33/10G01N33/02
Inventor 王战辉江海洋吴小平史为民徐飞王照鹏潘净茹王亚辉王熙
Owner 北京明日达科技发展有限责任公司
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