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Chemiluminesent immunoassay kit for detecting fipronil in tea

A chemiluminescence immunoassay and detection kit technology, applied in the field of immunological detection, can solve the problems of strong carcinogenicity and high toxicity of fipronil, and achieve the effects of simple instrument, reduction of use links, and guarantee of long-term effectiveness

Inactive Publication Date: 2019-05-14
ZHENJIANG YITE BIOTECH DEV CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Because fipronil has the characteristics of high toxicity and strong carcinogenicity

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] 1. Preparation of each component of the kit

[0041] (1) Preparation of fipronil hapten: acidify fipronil, react with sodium nitrite in a low-temperature environment without light at 4°C, and generate intermediates containing diazonium cations. The diazotized fipronil was used as a hapten for subsequent synthesis of immune antigens and coating antigens.

[0042] (2) Preparation of fipronil-bovine serum albumin (BSA) immunogen: Fipronil and bovine serum albumin (BSA) were coupled by diazotization method to obtain immune antigen.

[0043] Preparation of fipronil-oval serum albumin (OVA) coated antigen: coupling fipronil and egg serum albumin (OVA) by diazotization method to obtain coated antigen.

[0044]Preparation of fipronil-peroxidase-labeled antibody: For 6-8 week-old female BALB / c mice (body weight 18-20g), the high-dose immunization scheme is as follows: 160 µg fipronil-BSA and Mix an equal amount of complete Freund's adjuvant and inject subcutaneously. After 3 ...

Embodiment 2

[0073] The chemiluminescent immunoassay kit for detecting fipronil includes the following components:

[0074] (1) A 96-well opaque white microplate (8 wells x 12 strips) is coated with fipronil-mouse serum albumin conjugate, and is vacuum-sealed in an aluminum foil bag.

[0075] (2) 6 bottles of fipronil standard solution, the concentrations are:

[0076] 0 ng / mL, 0.02 ng / mL, 0.08 ng / mL, 0.32 ng / mL, 1.28 ng / mL, 5.12 ng / mL

[0077] (3) Fipronil-horseradish peroxidase labeled antibody solution.

[0078] (4) Luminescence substrate A solution (luminol and enhancer), luminescence substrate B solution (urea hydrogen peroxide).

[0079] (5) 2 times concentrated sample diluent. Before use, please dilute 1:1 (1 part concentrated diluent + 1 part deionized water) to become the working sample diluent, and the diluted working sample diluent is 0.05 mol / L, pH 7.4 Tris-HCl buffer.

[0080] (6) 20 times concentrated washing solution. Before use, please dilute at 1:19 (1 part of concent...

Embodiment 3

[0082] The chemiluminescent immunoassay kit for detecting fipronil includes the following components:

[0083] (1) A 96-well opaque white microplate (8 wells x 12 strips) is coated with fipronil-egg albumin conjugate, and is vacuum-sealed in an aluminum foil bag.

[0084] (2) 6 bottles of fipronil standard solution, the concentrations are:

[0085] 0 ng / mL, 0.02 ng / mL, 0.08 ng / mL, 0.32 ng / mL, 1.28 ng / mL, 5.12 ng / mL

[0086] (3) Fipronil-horseradish peroxidase labeled antibody solution.

[0087] (4) Luminescence substrate A solution (luminol and enhancer), luminescence substrate B solution (urea hydrogen peroxide).

[0088] (5) 2 times concentrated sample diluent. Before use, please dilute 1:1 (1 part of concentrated diluent + 1 part of deionized water) to become the working sample diluent, and the diluted working sample diluent is 0.05 mol / L, pH 7.4 glycine-HCl buffer.

[0089] (6) 20 times concentrated washing solution. Before use, please dilute at 1:19 (1 part of concen...

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Abstract

The invention provides a chemiluminesent immunoassay kit for detecting fipronil in tea, and belongs to the field of the immunological detection. The kit is composed of an opaque white ELISA plate coated with fipronil-carrier protein conjugate, fipronil standard substance, fipronil-peroxide enzyme-labeled antibody working solution, luminous substrate fluid, concentrated sample diluents, and concentrated washing solution. The fipronil-carrier protein conjugate is obtained by coupling the fipronil and the carrier protein through a mixed anhydride method or carbodiimide method; the concentrated washing solution contains 0.05% of tween-20. Compared with the traditional enzyme linked immunosorbent assay, the kit disclosed by the invention has higher sensitivity, and is low in detection time andlow in cost, and can be used for detecting the residual quantity of the fipronil in the tea sample.

Description

technical field [0001] The invention relates to a chemiluminescent immunoassay kit for detecting fipronil, which is used for detecting the content or residual amount of fipronil in animal-derived foods such as tea. It belongs to the field of immunological detection. Background technique [0002] Fipronil English common name is Fipronil (fipronil), trade name is Regent, is a broad-spectrum insecticide of phenylpyrazoles, which can kill fleas, mites, lice, aphids, leafhoppers , flies and coleoptera and other pests. It is mainly used to control various types of crop pests and sanitary pests in vegetables, rice, tobacco, cotton, animal husbandry, public health, storage supplies and the ground. [0003] Because fipronil is very toxic to bees and aquatic organisms and is not friendly to the environment, many countries have banned the use of this drug. According to the World Health Organization (WHO), ingesting high concentrations of fipronil for a sustained period of time can d...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/76G01N33/531
Inventor 洪霞杜霞立雯馨
Owner ZHENJIANG YITE BIOTECH DEV CO LTD
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