Study On Preparation Of ELISA Kit For Carbofuran And Detoxic Function Of Anti-carbofuran Antibody

Two kinds of haptens 6-{[(2,3-Dihydro-2,2-Dimethyl-7-benzofuranyloxy)carbonyl] -amino}hexanoicAcid(BFNH)and4-{[(2,3-Dihydro-2,2-Dimethyl-7-benzofuranyloxy) carbonyl]-amino}butanoic Acid(BFNB) has been synthesized by chemical way. Then they were covalently conjugated to bovine serum albumin (BSA) or ovalbumin(OVA) to prepare antigens(immunogens) and coating antigen The conjugates were identified and their molecular ration of hapten to carrier protein were determined by UV spectrophotometry. Rabbit were immunized with synthetic antigens and then two kinds of antisera specific to carbofuran were obtained by using BFNH-BSA, BFNB-BSA as immunogens. The titers of the antisera were ranged froml: 16,000 to 1:128,000 and the purified antibody's titers were ranged from 1:500,000 to 1:4,000,000, respectively. The cross reactives of anti-BFNB-BSA serum to carbofuran-phenol, carbosulfan were 4.00% and 6.19%, other carbamates were less than 0.44%, three organophosphorus pesticide were less 0.12%.The enzyme tagged antibody(E-Ab) was prepared by coupling horseradish peroxidase(HRP). The enzyme conjugate maintained both immunological activity and enzyme activity and its liter was 1:4000.Two ELISA kits(Enzyme-linked Immunosorbent Assay kit) including indirect ELISA and E-Ab direct ELISA, have been established to detect the residue of carbofuran in specimens of water > soiK vegetable and food poisoning samples. The in-ELISA kit's linear detection was ranged from 0.005 to 10.00 mg/L, and the E-Ab ELISA kit's linear detection was ranged from 0.01 to 10.00 mg/L. The detection limits were 0.005mg/L and O.Olmg/L for two kinds of kits, respectively. The mean within-assay > between-assay and total-assay variability were less than 8.0%. The average recoveries of carbofuran added to sample were above 89.62% except for vomitus sample. The validity of kits was six months or more.Since specific antibodies could be generated to target antigens, the experiment study was carried out on application of antibody in medical way to prevent inhibition in mice roisonned by carbofuran. The result indicated that the toxic effect of carbofuran on mice acetylcholine esterase was significantly reduced by the antibody, and the mice were recovered as normal.