Phosphonic acid (trans-4-phosphono-2-butenic acid; TPB) was used as a generic hapten in order to generate broad specificity antibodies against a group of organophosphorus pesticides. The polyclonal antiserum showed, in an indirect enzyme-linked immunosorbent assay (ELISA) format, preferential binding toward pesticides containing unsaturated diethyl-phosphate functionalities rather than the equivalent thiophosphate or dimethyl structures. The level of detection in the ELISA using a heterologous system was investigated and showed a 20-fold improvement when a conjugate for which the antibody had lower affinity was immobilized on the plate. Biosensor assays using parathion as a standard indicated that the antibody had a relatively high dissociation rate, and reproducible cycles of regeneration were achieved. The potential for using TPB as a generic hapten is discussed.

A rabbit polyclonal antiserum and two murine monoclonal antibodies recognizing the organophosphorus pesticide chlorpyrifos-ethyl were produced. The two hybridoma cell lines were then used as sources of immunoglobulin genes for the generation of recombinant scFv antibodies in Escherichia coli. The two scFvs showed either similar or improved limits of detection in an ELISA when compared with the monoclonal antibodies. Cross-reactivity studies showed that all of the antibodies were specific toward the chlorinated aromatic ring. Furthermore, scFv gene sequences were linked directly to sequences coding for either a c-Myc tag, a His-tag, or alkaline phosphatase. The fusion products generated were functional, and their properties were determined. The problems associated with producing scFvs and scFv derivatives for detection of pesticide residues from hybridoma are addressed and discussed.