Detection and quantification of the organophosphate insecticide paraoxon by competitive inhibition enzyme immunoassay

Abstract

A competitive inhibition enzyme immunoassay (EIA) was developed to detect and quantify levels of the organophosphate insecticide paraoxon in body fluids. Protein-conjugated paraoxon served as an immunogen for the production of rabbit heteroantiserum, from which affinity purified IgG anti-paraoxon antibodies were isolated using a heterologous protein-paraoxon-conjugated immunoabsorbent. In the competitive inhibition EIA a standard curve was generated for the inhibition of binding of anti-paraoxon IgG to a solid-phase bound heterologous protein-paraoxon conjugate by various concentrations of free paraoxon. Binding was proportionate to the color change of an appropriate substrate generated by an enzyme-conjugated second antibody specific for the rabbit IgG anti-paraoxon. The assay detected paraoxon levels as low as 10 −10M (28 pg/ml) in buffer, and serum paraoxon levels as low as 10 −9M. In addition to its sensitivity, this technique is ideally suited to the simultaneous processing of large numbers of samples in less than 2 hr. The competitive inhibition EIA is cost effective and should facilitate environmental surveillance using sentinel animals, expand laboratory toxicology studies, and improve clinical detection capabilities.