The Research On Enzyme-Linked Immunosorbent Assay For Fenvalerate And Fenpropathrin

The using of pyrethroid widely brings a lot of problems, such as the safety of agricultural products and the pollution. For the research of immunochemistry characteristic of pyrethroid insecticides, we choose fenvalerate and fenpropathrin as being studied objects. We have developed an ic-ELIS A for fenvalerate.Two haptens were designed and synthesized for fenvalerate from 2-(4-chlorophenyl)-3-methyl-butyric acid and a-cyano-3-phenoxyhenzyl-benzyl- alcohol, the immunizing hapten FVb and the coating hapten FVa. The hapten FVb was conjugated to bovine serum albumin(BSA) with the CDI method to form immuno-antigen; the hapten FVa was conjugated to ovalbumin(OVA) with mixed anhydride method to form the coating antigen. The ultraviolet absorption spectra shows that the reactions succeed. And the link ratios were 35:1 and 10:1. The New Zealand rabbits were immunized by the conjugate FVb-BSA, and titres of anti- fenvalerate serum (1.28×10⁴) was determined by non-competitive indirect enzyme- linked immunosorbent assay. The antibody was got via the purification of the serum. The antibody was frozen to be freeze-dried powder. The titre(1×10⁵) of the freeze-dried powder was also determined by non-competitive indirect enzyme- linked immunosorbent assay. After optimization of the ic-ELISA, such as pH values, inonic strengths, concentrations of methanol, pH 7.5, PBS of 0.3 M Na⁺, and thirty percent methanol were determined to optimum assay condition. The IC₅₀ for fenvalerate was 1.19 mg/L, and lower detection limit(LDL) was 0.017mg/L. Pyrethroids, such as fenpropathrin, cyhalothrin, permethrin, cypermethrin, deltamethrin and bifenthrin, did not cross-react significant in this assay. When the fenvalerate was added in the distill water, the recovery rate was 83.5%～110.2%.Two haptens were designed and synthesized for fenpropathrin from 2,2,3,3-tenramethyl cyclopropane carboxy andα-cyano-3-phenoxyhenzyl-benzylalcohol, the immunizing hapten FPb and the coating hapten FPa. The hapten FPb was conjugated to bovine serum albumin(BSA) with the CDI method to form immuno-antigen; the hapten FPa was conjugated to ovalbumin(OVA) with mixed anhydride method to form the coating antigen. The ultraviolet absorption spectra shows that the reactions succeed. And the link ratios were 38:1 and 15:1. The New Zealand rabbits were immunized by the conjugate FPb-BSA, and titres of anti- fenpropathrin serum (1.28×10⁵) was determined by non-competitive indirect enzyme-linked immuno- sorbent assay. The antibody was got via the purification of the serum. The antibody was frozen to be freeze-dried powder. The titre(5.12×10⁵) of the freeze-dried powder was also determined by non-competitive indirect enzyme- linked immunosorbent assay. The antibody was not sensitive to fenpropathrin, as well as the other pyrethroids. But the IC₅₀ for FPb was 9.33 mg/L. There may be three reasons: the hapten design, the hapten synthesis, and the immunity operation.The titre of the antibody for fenpropathrin was higher than the antibody's for fenvalerate. When the antibody was used to detect the pesticide, the result is reversed. The hapten synthesis method, the emulsification and the immune scheme are all the same. But the result is different. It needs more research to confirm the reason.