Robust and Sensitive Monoclonal Enzyme-Linked Immunosorbent Assay for the Herbicide Molinate

Caterina Rufo, Bruce D. Hammock, Shirley J. Gee, Jerold A Last, Gualberto González-Sapienza

Research output: Contribution to journalArticle

14 Scopus citations

Abstract

This paper reports on the generation of monoclonal antibodies and the development of a new enzyme-linked immunosorbent assay (ELISA) for the detection of molinate (S-ethyl hexahydroazepine-1-carbothioate). Hybridoma cells were generated using spleen and lymph node cells from a mouse immunized with S-2-carboxyethyl hexahydroazepine-1-carbothioate conjugated to keyhole limpet hemocyanin. After screening with a competitive ELISA, two monoclonal antibodies, mAbs 16C11 and 14D7, with IC50 values of 82 ± 2 and 173 ± 8 ng/mL, respectively, were selected. MAb 16C11 can detect molinate concentrations of 1 ng/mL with no cross-reactivity to any other thiocarbamate pesticides; however, it was susceptible to the presence of organic solvents and to variation in buffer ionic strength. MAb 14D7 tolerated concentrations up to 5% of propylene glycol and 12.5% of acetonitrile in the assay buffer. The sensitivity of mAb 14D7 was further improved by decreasing the amount of coating antigen in the ELISA; the final inhibition assay showed an IC50 of 69.2 ± 1.4 ng/mL. In summary, mAb14D7 provided a more sensitive and robust assay, as compared with previous polyclonal antibody-based assays, with the additional advantage of being based upon a consistent and unlimited source of a defined reagent.

Original languageEnglish (US)
Pages (from-to)182-187
Number of pages6
JournalJournal of Agricultural and Food Chemistry
Volume52
Issue number2
DOIs
StatePublished - Jan 28 2004

Keywords

  • ELISA
  • Molinate
  • Monoclonal antibody
  • Thiocarbamate herbicide

ASJC Scopus subject areas

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

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