Study On Fluorescence Polarization Immunoassay And Multi-Residue Elisa For Organophosphorus Pesticide

The aim of the research is development of the flurescence polization immunoassays (FPIAs) and the multi-residue enzyme linked immunosorbant assays (ELISAs) of organophosphorus pesticides (OPs). the 3 kinds of haptens were synthesized. The 3 antisera, which were anti- O,O-diethyl phosphate, anti- aromatic O,O-diethyl phosphorothioate and anti- alkylate O,O-diethyl phosphorothioate, were produced by immunogen conjugated between hapten and bovin serum alburmin (BSA). The 2 FPIAs and 3 ELISAs for multi-residue were developed on the basis of the antisera. The analytical methods will provide the technological bases for OPs residue determination in environment.The results are showed as following:1. DEP was adopted as hapten and conjugated with BSA with active ester method. The immungen (DEP-BSA), coatingen (DEP-OVA) were the conjugator of hapten with BSA and OVA, respectively. The anti- O,O-diethyl phosphate antisera were obtained by DEP-BSA. And the titers of the 3 antisera were detected by ELISA. And the titers of antisera A, B and D were 1︰10000,1︰5000 and 1︰20000, respectively.The ELISA was developed with antiserum D. And the several kinds of influence factors were discussed in the research. The result was that the 2μg/mL coatingen concentration, coating overnight at 4℃and pH=7.0 reaction circumstances was the optimal condition for ELISA.The antisera were affinitive with the O,O-diethyl phosphate pesticides by affinitive experiment. The multi-residue ELISA was developed using the antiserum D. And the recoveries of parathion, pirimiphos and dichlorvos in the water samples were from 106.0% to 110.0%.2. The FPIA for parathion was developed on the basis of anti-parathion antibodies A, D and the tracer of PM-B-EDF. The recoveries of parathion in the water samples ranged from 104.000% to 108.280%, with the LOD of 0.18μg/mL.The FPIA for azinphos-methyl was developed on the basis of anti- azinphos-methyl antibody MAb-Azi-110 and the tracer of MBP-EDF 2. The recoveries of azinphos-methyl in the water samples ranged from 96.0% to 120.0%, with the LOD of 0.148μg/mL.3. The hapten of ArOP was obtained by reducing parathion. The immungen (ArOP-BSA) and coatingen (ArOP-OVA) were got from conjugator of hapten with BSA and OVA, respectively. The anti-aromatic O,O-diethyl phosphorothioate antisera were abtained by ArOP-BSA. And the titers of the 3 antisera were detected by ELISA. And the titers of 3 antisera were 1︰40000,1︰80000 and 1︰40000, respectively.The antisera were affinitive with the aromatic O,O-diethyl phosphorothioate pesticides by affinitive experiment. The multi-residue ELISA was developed using the antiserum 2. And the recoveries of parathion and diazinon in the water samples were from 96.0% to 106.4%.4. The hapten of DENP was obtained by reaction between O,O-diethylthiophosphoryl chloride and 4-animobutycarboxyl. The immungen (DENP-BSA) and coatingen (DENP-OVA) were got from conjugator of hapten with BSA and OVA, respectively. The anti- alkylate O,O-diethyl phosphorothioate antiserum was obtained by DENP-BSA. And the titer of antiserum 1︰50000 was detected by ELISA.The antisera were affinitive with the alkylate O,O-diethyl phosphorothioate pesticides by affinitive experiment. The multi-residue ELISA was developed using the antiserum. And the recoveries of demeton S and phorate in the water samples were from 108.0% to 112.0%, with the LOD of 0.043μg/mL and 0.02μg/mL, respectively.