Technology Research On Immunoassay Of Dimethachlon

Dimethachlon is a dicarboximides fungicide which is effective for Sclerotinia sclerotiorum,Alternaria alternata,Rhizoctonia cerealis and so on. It is necessary to develop a sensitive,fast and economical screening method to monitor levels of dimethachlon residue. Immunoassay is an important tool for quick analysis and detection of pesticide and its derivatives with high sensitivity and high specificity. In this paper, the immunoassay technology of dimethachlon was studied. The primary research results were as follows:1. To produce a novel type of immunogen susceptible to raise antibodies, the dimethachlon-2-hemigiutarate (hapten) mimicking the analyte, dimethachlon, was synthesized and conjugated with the carrier proteins bovine serum albumin (BSA) and ovalbumin (OVA) by active ester method (DCC). The immunogen and the coated antigen were identified by UV spectrophotometry and SDS-PAGE.2. Polyclonal antibodies were raised through immunizing New Zealand white rabbits with hapten-BSA . The titers of antiserum had reached 1:512000. The optimal working concentration were optimized by ELISA as follows: the dilution ratio of coated antigen was 1:1000 (2μg/mL), the dilution ratio of antiserum was 1:128000.3. An indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) for dimethachlon was developed, and its suitability for the determination of this analyte in spiked tobacco samples was studied. The IC-ELISA showed an I₅₀ value of 2.83μg /mL with a detection limit of 0.011μg/mL and a response range for dimethachlon between 0.044-25.00μg/mL. The optimal detection conditions were as follows: the methanol content in samples was not more than 10%; the pH of PBS Buffer was 7.4 for higher sensitivity of ELISA. In spiked tobacco samples, recoveries determined by IC-ELISA and HPLC were in the range of 65.58%-80.88% and 64.19%-73.05%, respectively. A good correlation between dimethachlon concentration measured by ELISA and HPLC was obtained from linear regression analysis (R²=0.9965, y=0.8934x + 1.2089). The IC-ELISA for dimethachlon could be used for quick test and screen of dimethachlon residues.