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Fipronil immune detecting method

An immunological detection method and fipronil technology, which is applied in the direction of testing food, measuring devices, biological testing, etc., can solve the problems of complex detection process, and achieve the effects of simple operation, short testing process and strong fluidity.

Inactive Publication Date: 2003-04-30
江苏省农业科学院植物保护研究所
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The average recovery rate is 85.4-95.8%, and the coefficient of variation is 2.3-12.0%. The detection result generally takes two days, and the detection process is complicated

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1: Preparation method of coated antigen.

[0020] First, the nitrile group on the pyrazole ring of fipronil is transformed into a carboxyl group, and the carboxyl group is coupled with the amino group of ovalbumin by the activated ester method to obtain a fipronil-ovalbumin conjugate, and obtain a coated antigen diluted antigen.

Embodiment 2

[0021] Embodiment 2: Solid-phase indirect detection steps.

[0022] 1. Coating: Add 100ul of coating solution to each well of a 96 microwell plate, and incubate at 4°C overnight or at 37°C for 2 hours;

[0023] 2. Blocking: Take out the 96-microwell plate, discard the coating solution, wash with PBST three times, spin dry, and add 200ul of blocking solution to each well, and incubate at 37°C for 1 hour;

[0024] 3. Adding samples: Take out the ELISA plate, wash it three times with PBST, dry it, remove the suspended impurities from the sample, mix it with an equal volume of antibody mixture, inject it into a 96 microwell plate as shown in Table 1, 100ul per well, At the same time, microplate A 7 , B 7＝ 、C 7 Well add 100ul of PBS, A 8 , B 8 、C 8 Add 50ul of antibody and 50ul of PBS, incubate at 37°C for 2 hours;

[0025] 4. Add enzyme-labeled SPA: take out the enzyme-labeled plate, wash with PBST, spin dry, add 100ul enzyme-labeled SPA diluent to each well, and incubate a...

Embodiment 3

[0031] Embodiment 3: Solid phase direct detection procedure.

[0032] 1. Coating: add 100ul of coating antigen diluent to each well of a 96 microwell plate, and incubate overnight at 4°C or 37°C for 2 hours;

[0033] 2. Blocking: Take out the 96-microwell plate, discard the coating solution, wash with PBST three times, spin dry, and add 200ul of blocking solution to each well, and incubate at 37°C for 1 hour;

[0034] 3. Adding samples: Take out the ELISA plate, wash it three times with PBST, dry it, extract the sample with acetone and water at a ratio of 95:5, purify and concentrate the extract, mix the treated sample with an equal volume of antibody mixture, Inject into 96 microwell plate as shown in Table 1, 100ul per well, meanwhile, ELISA plate A 7 , B 7＝ 、C 7 Well add 100ul of PBS, A 8 , B 8 、C 8 Add 50ul of antibody and 50ul of PBS, and incubate at 37°C for 2 hours.

[0035] 4. Add substrate solution: take out the microplate, wash it three times with PBST, and sh...

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PUM

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Abstract

The method includes the following procedures: incubating the diluent of the peridium antigen. Adding a confining liquid and incubating. Mixing the processed specimen with the enzyme-marked antibody and incubating. Taking out the enzyme-marked board, adding the substrate fluid and incubating. Adding sulphuric acid solution to end the reaction. 450mm wavelength, determining the OD value and detecting the fipronil content in the specimen. The invention provides the features of easy of operation, the short detecting procedure. The invention can be utilized in the application and research area such as the environment control, the food industry and pesticide. It is particularly suitable for the grass root and the markets monitors section.

Description

Technical field: [0001] The invention relates to a fipronil immunoassay method. Background technique: [0002] Fipronil is a new type of benzopyrazole insecticide, which has a good application prospect in my country, but studies have shown that it is a chronic neurotoxicity, and it can cause thyroid cancer in mice at an oral dose of 300ppm. Class C carcinogens, especially its slow metabolism in the environment, two metabolites are more than 10 times more toxic to mammals than parent pesticides, and have enrichment in the fat of organisms, especially the pesticides on Aquatic crustaceans are highly toxic, and are highly toxic to bees. They are a kind of pesticide with relatively toxic and negative effects on the ecological environment. It is necessary to strengthen the supervision of this pesticide after application. [0003] He Yibing disclosed a gas chromatography analysis method in the third issue of "Scientific Management of Pesticides" in 2000. The requirements for instru...

Claims

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Application Information

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IPC IPC(8): G01N33/02G01N33/53G01N33/535G01N33/543G01N33/577
Inventor 刘贤金颜春荣董键余向阳
Owner 江苏省农业科学院植物保护研究所
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