The Research On Enzyme-Linked Immunosorbent Assay For S-Bioallethrin

About25%of current-use pesticides are chiral. Almost all of the pyrethroids are chiral pesticides. Due to their stereoselective interactions in biological systems, enantiomers often behave drastically differently in their toxicity and environmental fate. Therefore, in order to reduce pesticide use and protect the environment from unintended effects, it is greatly advantageous to study the determination assay of chiral pesticides.Immunosorbent assay provide a highly sensitive, selective, simple and field-analytical method for the detection of pesticide residues. Indirect competitive enzyme-linked immunosorbent assays (ic-ELISA) and Enhanced Chemiluminescence Enzyme Linked Immunoassay (ECL-ELISA) were developed for the quantitative detection of S-bioallethrin in the environmental and agricultural samples.A hapten of S-bioallethrin was synthesized. The hapten-bovine serum albumin (BSA) conjugate was used as an immunogen, while the hapten-ovalbumin (OVA) conjugate was applied as a coating antigen. A sensitive indirect competitive ELISA based on polyclonal antibody for detection of S-bioallethrin was developed. The conjugates were confirmed by UV spectra, the conjugation molar ratio is32:1and10:1for the immunogen and coating antigen. Polyclonal antibodies was generated by immunizing male New Zealand rabbits with the immunogen, the titers of the antibodies were8×105. The optimal concentrations of coating antigen and antiserum were determined by a checkerboard titration assay. The inhibition standard curve of S-bioallethrin was obtained under the optimised conditions (40%methanol,0.5mol/L Na+, pH6.5). The half-maximal inhibition concentration (IC50) was0.089mg/L and the limit of detection (LOD)(IC10) was0.0025mg/L. Very low cross-reactivity was found for some structurally related compounds except ES-bioallethrin、 Rich-trans-allethrin、allethrin, demonstrating that Abs have highly stereoselectivity. After using ECL-ELISA, IC50was further improved3.56fold to0.025mg/L and the limit of detection(LOD)(IC10) was further improved2.5fold to0.001mg/L。At spiked different levels (0.05-0.5mg/L for ELISA and0.01-0.1mg/L for ECL-ELISA) recoveries of S-bioallethrin in tap water, river water, distilled water and dust samples were 90.7%-111.7%and78.9%-100.7%with CV of1.2%-6.5%and1.2%-3.9%, respectively. The ELISA and ECL-ELISA for spiked samples was confirmed by gas chromatography/electron capture detection (GC-ECD) with high correlation coefficient of0.9882and0.998(n=12). The pAb-based ELISA and ECL-ELISA could be a high sensitivity, simplicity, rapidity and lower expenses method for S-bioallethrin analysis in environmental samples.