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Enzyme linked immunoassay kit for detecting abamectin and preparation method and detection method of enzyme linked immunoassay kit

A technology of abamectin enzyme and immune reagent, applied in the detection of abamectin ELISA kit and its preparation field, which can solve the threat to human life, cannot detect abamectin, and cannot detect abamectin element and other issues

Inactive Publication Date: 2015-01-14
贵州勤邦食品安全科学技术有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] It can be seen that ELISA has been applied to a large extent in the detection of abamectin, and it also enables low-concentration abamectin drugs to be detected from food, thereby ensuring food safety But, despite this, the enzyme-linked immunosorbent assay in the above-mentioned prior art still can not relatively ideally detect the abamectin in the food, and what is disclosed is also all the detection in the meat food field, so that the abamectin in the food The detection result of the content of vermectin is inaccurate, therefore has brought threat to human life; This is mainly because, the sensitivity of the ELISA detection avermectin in the prior art and the specificity of kit are higher; Furthermore, the content of avermectin in it cannot be detected, but the reduction product of avermectin in food and other similar mycocins are detected

Method used

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  • Enzyme linked immunoassay kit for detecting abamectin and preparation method and detection method of enzyme linked immunoassay kit
  • Enzyme linked immunoassay kit for detecting abamectin and preparation method and detection method of enzyme linked immunoassay kit
  • Enzyme linked immunoassay kit for detecting abamectin and preparation method and detection method of enzyme linked immunoassay kit

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Experimental program
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Effect test

Embodiment 1

[0076] A preparation method for detecting abamectin ELISA kit, comprising the following steps:

[0077] (1) Immunogen preparation: abamectin and bovine serum albumin are obtained by coupling according to the weight ratio of 24:1, and are ready to use;

[0078] (2) Coating raw preparation: combine avermectin and bovine serum albumin according to the weight ratio of 21:1 and then couple them, and then use them;

[0079] (3) Preparation of monoclonal antibody: Take the immunogen obtained in step 1) to immunize female BALB / c mice, which are 8-12 weeks old; the specific immunization method is: when first immunizing, each mouse is immunized with 100 μg Mix AVM-BSA with the same amount of FCA to make an emulsifier, and inject it subcutaneously at multiple points on the back of the neck; for the second immunization, each mouse mixes 100 μg of AVM-BSA with the same amount of FICA to make an emulsifier, and inject it at the back of the neck Subcutaneous injection at multiple points; th...

Embodiment 2

[0086] An ELISA kit for detecting abamectin, including an immunogen, a coating source, a monoclonal antibody, an enzyme-labeled secondary antibody, and a TMB substrate chromogenic solution; wherein the immunogen is obtained by coupling abamectin with BSA , the binding ratio is 24:1; the coating is obtained by coupling avermectin and BSA, and the binding ratio is 21:1; the immunoglobulin subclass of the monoclonal antibody is IgG1, the molecular weight is 165.7KDa, the number of chromosomes There are 89 to 94 strips, and the affinity constant is 1.80×1010M-1; the TMB substrate chromogenic solution is composed of A solution and B solution, and A solution is hydrogen peroxide buffered by diluting hydrogen peroxide with distilled water at 1:10000 times. Solution, solution B is a solution obtained by dissolving 200 mg of tetramethylbenzidine (TMB) in a citric acid-phosphate buffer solution with a volume of 100 ml, a molar concentration of 100 mmol / L, and a pH of 5.0;

[0087] The d...

Embodiment 3

[0091] A method for detecting avermectin enzyme-linked immunoimmunoassay kits for abamectin detection. The sample solution is mixed with the coating original, and then mixed with the monoclonal antibody solution according to the volume ratio of 20:80, and then reacted After 30-60min, adjust the temperature to 35-38°C, then add the enzyme-labeled secondary antibody to it, react for 30min, then add TMB substrate chromogenic solution at constant temperature, wait for it to react for 15min, and then use it with the indirect competition ELISA The standard curve of the kit is compared to obtain the content of abamectin.

[0092] The dilution factor of the coating agent is 1:3000, and it is diluted with distilled water.

[0093] The enzyme-labeled secondary antibody has a dilution factor of 1:1000 and is diluted with distilled water.

[0094] The monoclonal antibody has a dilution factor of 1::40000 and is diluted with distilled water.

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Abstract

The invention relates to the technical field of abamectin detection, and in particular relates to an enzyme linked immunoassay kit for detecting abamectin and a preparation method and a detection method of the enzyme linked immunoassay kit. The enzyme linked immunoassay kit comprises an immunogen, a coating antigen, a monoclonal antibody, an enzyme-labeled second antibody and TMB substrate developing liquid, wherein the immunogen is obtained by coupling abamectin and BSA according to a combination ratio of 24 to 1; the coating antigen is obtained by coupling abamectin and BSA according to a combination ratio of 21 to 1; the immunoglobulin subclass of the monoclonal antibody is IgG1, the molecular weight is 165.7KDa, the quantity of chromosomes is 89-94, and the affinity constant is 1.80*10<10>M<-1>; the TMB substrate developing liquid consists of liquid A and liquid B. The enzyme linked immunoassay kit has the advantages of high stability, excellent specificity, long expiration date, simple preparation technology, relatively high sensitivity, relatively high accurate abamectin detection and simplicity in use.

Description

technical field [0001] The invention relates to the technical field of abamectin detection, in particular to an ELISA kit for detecting abamectin, a preparation method and a detection method thereof. Background technique [0002] Abamectin drugs are macrolide antibiotics. The commercialized avermectin is a compound species of eight very similar chemical structures, the most active mixture of the two homologous systems. The reduction product is ivermectin, another important antiparasitic drug. Abamectin has a strong inhibitory effect on the vast majority of parasitic nematodes and arthropods with important economic value. Its insecticidal activity and insecticidal spectrum are epoch-making, and its mechanism of action is unique, easy to use, and It is regarded as a major breakthrough in the research of antiparasitic drugs in the past ten years. At present, abamectins have begun large-scale production and application. [0003] However, the excessive use of abamectin drug ma...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/94G01N33/535
CPCG01N33/535G01N33/577
Inventor 罗贵昆谢体波蒲小容易重任金帮明刘天雷何方洋
Owner 贵州勤邦食品安全科学技术有限公司
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