Development of an enzyme-linked immunosorbent assay for the detection of the organophosphorus insecticide acephate

Jae Koo Lee, Ki Chang Ahn, Donald W. Stoutamire, Shirley J. Gee, Bruce D. Hammock

Research output: Contribution to journalArticlepeer-review

41 Scopus citations


A competitive indirect enzyme-linked immunosorbent assay (ciELISA) for the organophosphorus insecticide acephate, O,S-dimethyl acetylphosphoramidothioate, was developed using a polyclonal antibody. Five different haptens mimicking the analyte were synthesized and conjugated with the carrier proteins bovine serum albumin (BSA) and keyhole limpet hemocyanin (KLH) by the N-hydroxysuccinimide active ester and diazotization methods. Polyclonal antibodies raised against hapten - KLH conjugates in rabbits and hapten - BSA conjugates as coating antigens were screened and selected for the assay in the homologous and heterologous ELISA systems. The effects of various assay conditions such as detergent, organic solvents, pH, and preincubation of the mixture of the polyclonal antibody and the analyte on the sensitivity were evaluated. The IC50 value for acephate was 25 ng/mL in an optimized heterologous system using hapten-4 - BSA as a coating antigen and a polyclonal antibody no. 8377 against hapten-1 - KLH, showing the detection range of 5-140 ng/mL and the lowest detection limit of 2 ng/mL. The cross-reactivities of the structurally related organophosphorus insecticides, including the major metabolite of the analyte, methamidophos, were less than 1%. Recoveries from the analyte-fortified tap water, mulberry leaves, and lettuce samples in the assay were in the range of 72-121% by simple extraction, concentration, and dilution. These results indicate that the ELISA could be a convenient and supplemental analytical tool for monitoring acephate residues in environmental and agricultural samples.

Original languageEnglish (US)
Pages (from-to)3695-3703
Number of pages9
JournalJournal of Agricultural and Food Chemistry
Issue number13
StatePublished - Jun 18 2003


  • Acephate
  • Organophosphorus insecticide
  • Polyclonal antibody
  • Residue analysis

ASJC Scopus subject areas

  • Agricultural and Biological Sciences (miscellaneous)
  • Food Science
  • Chemistry (miscellaneous)


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