A monoclonal antibody-based ELISA for the analysis of azinphos-methyl in fruit juices

A monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA) was developed for the quantification of azinphos-methyl (O,O-dimethylS-[(4-oxo-1,2,3-benzotriazin-3-yl)methyl]phosphorodithioate). The competitive ELISA was performed in the antibody-coated format using a homologous peroxidase-hapten conjugate as enzyme tracer. For standards, the I50 value of the ELISA was around 3 ng ml−1 with a detection limit of 0.4 ng ml−1. On the basis of previous crossreactivity studies, multianalyte standards containing the three pesticides recognized by the antibody (azinphos-methyl, azinphos-ethyl, and phosmet) were assayed. Results showed that the total content of these analytes can be accurately determined using a unique azinphos-methyl standard curve, as long as the three compounds are in equimolar amounts. The suitability of the ELISA for pesticide quantification in orange and apple juices was also studied. The practical detection limit in juice samples was 100 ng ml−1. Therefore, samples were spiked with azinphos-methyl in the 100–1000 ng ml−1 range and directly analyzed without any pretreatment other than a 1100 dilution in PBS to minimize matrix effects. Under these conditions, good accuracy and precision were obtained, with mean recoveries of 122.7% and 110.5%, and mean coefficients of variation of 14.5% and 15.1%, for orange and apple juices, respectively. No false positives were found.