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%.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

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

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

Example Embodiment

[0021] Example 2: Solid phase indirect detection step.

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

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

[0024] 3. Add sample: Take out the ELISA plate, wash it with PBST three times, spin dry, remove the suspended impurities, mix with an equal volume of antibody mixture, and pour into a 96-well plate as shown in Table 1, 100ul per well, At the same time, the microplate A 7 , B 7＝ , C 7 Add 100ul PBS to the well, A 8 , B 8 , C 8 Add 50ul of antibody and 50ul of PBS and 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 of the enzyme-labeled SPA diluent to each well, and incubate at 37°C for 45 minutes;

[0026] 5. Add s...

Example Embodiment

[0031] Example 3: Direct solid phase detection step.

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

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

[0034] 3. Add sample: Take out the ELISA plate, wash it three times with PBST, spin dry, extract the sample with acetone and water at 95:5, purify and concentrate the extract, and mix the processed sample with an equal volume of antibody mixture. Pour into 96 micro-well plate as shown in Table 1, 100ul per well, meanwhile, ELISA plate A 7 , B 7＝ , C 7 Add 100ul PBS to the well, 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 ELISA plate, wash three times with PBST, and spin dry. Add freshly prepared substrate...

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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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