Immunoassay For Typical Pesticide And Veterinary Drugs Residue Basedon Magnetic Nanoparticals

Over the years, food safety problems caused by pesticide and veterinary drug residues happened regularly both in China and abroad. Development of reliable, fast and sensitive analysis of residue in food has become urgent research. Recently, immunoassay technique has been a common technique and research focus in residue detection because of its rapid reaction. However, the existing immunoassay can not meet the needs of actual testing because of the limits of sensitivity and class-specificity and be limited with the unstable enzyme used. The research is to introduce magnetic nanoparticals (MNPs) into immunoassay and to make the first study of developing immunoassay for typical pesticide and veterinary drugs residue based on MNPs. The research will provide the technological bases for improving sensitivity of detection,solving classic-specificity issues in multiresidue analysis and being a substitute for enzyme.(1) Fe3O4 magnetic nanoparticals with diameter 10nm were preparaed by coprecipitation and modified with 3-aminopropyl triethoxysilane (APTES), then coated with immune molecules at the concentration of 1:1000. Pictures of transmission electron microscopy(TEM) show that particles modified with APTES haven't changed the diameter so much, and the MNPs solution has performed good stability with a more than 1.5h half-settle time and a 57% of coating rate.(2) MNPs'enzyme-like activity and signal amplification were studied. The best activity was based on the optimality condition: 200μL 0.6% (w/v) TMB/DMSO, 20μL 75% H2O2, 20℃, pH=4 and 6min's acting time.(3) DC-ELISA for clenbuterol based on Fe3O4's enzyme-like activity was developed.Results showed that enzyme used in DC-ELISA can be substituted by Fe3O4 without LOD reduction. The LOD and linear range were 0.081 ng/mL and 0.177~29.3 ng/mL, the cross-reaction rate and recovery rate have met the requirements.(4) ELISA(IC-ELISA, MFA-ELISA and MSA-ELISA) for OPs have been developed. Comparing with IC-ELISA, MFA-ELISA has made the reduction range of LOD at 72.9% (chlopyrifos) and 76.1% (quinalphos) based on the optimality condition: 5% of methanol in solution, pH=7.4, the antibody dilution 1:1600. And the cross-reaction rate to OPs has been improved with the detection results of IC50 reduction range from 68.3~95.6%.(5) Results of the cross-reaction and recovery to chlopyrifos and other 14 kinds of pesticides show that mp-ELISA gets a higher sensitivity and class-specificity with the mean cross-reaction 34% which is 127% higher than IC-ELISA. And the recovery rate of MFA-ELISA is 78-107% to chlopyrifos, 77-110% to quinalphos, which has met the requirements.(6) The comparation of IC-ELISA and MFA-ELISA for clenbuterol has shown that MFA-ELISA got a high sensitivity with a reduction range of IC50 at 28%. Recovery from pork samples were 82.3~115.1%(IC-ELISA) and 83.1~105.6%(MFA-ELISA).(7) Immunochromatographic assay strips based on MNPs have been developed, which got a similar sensitivity of traditional strips and advantages of lower cost,simply producing & using and ability of semi-quantitative.