Synthesis of derivatives and production of antiserum for class specific detection of pyrethroids by indirect ELISA

Abstract

Two series of haptens including 3-phenoxybenzoic acid (PBA) and 3-(2-chloro-3, 3,3-trifluoroprop-1-enyl)-2,2-dimethylcyclo-propanecarboxylic acid (CF₃MPA) were used to prepare immunogens through attachment of 4-C or 6-C handles. Class selective antibodies were produced by immunising rabbits. Ab502 showed the highest reactivity towards tau-fluvalinate (IC₅₀ 1.3 ng mL⁻¹), λ-cyhalothrin (IC₅₀ 2.3 ng mL⁻¹), cyfluthrin (IC₅₀ 2.2 ng mL⁻¹) and fenpropathrin (IC₅₀ 18.5 ng mL⁻¹) among the antibodies in a competitive ELISA. The effects of methanol, pH and salt concentration were optimised for maximum efficiency of the ELISA (Enzyme-Linked ImmunoSorbent Assay). Ab502 (1:80000)/2-OVA-1(0.2 µg mL⁻¹) was chosen for ELISA optimisation. Finally, 0.05 M phosphate buffered saline (PBS) at pH 6.5 containing 30% methanol (v/v) was used to dilute the standards. Target analytes in honey samples were extracted with ethyl acetate by sonication. The samples were spiked with three different concentrations of each compound (tau-fluvalinate, 0.5 ng g⁻¹, 3 ng g⁻¹, 12 ng g⁻¹; λ-cyhalothrin and cyfluthrin 1 ng g⁻¹, 5 ng g⁻¹, 65 ng g⁻¹). The recoveries were 36–59% at the lowest spiking concentration and 61–81% at the higher concentration. This assay might be useful to screen pyrethroid residues in honey or other matrix.

Keywords:

• pyrethroid
• ELISA
• class selective antibody
• tau-fluvalinate

Acknowledgements

We appreciate the help from He Yong and Feng Jing of Wanger Co. Ltd., China on feeding rabbits and animal immunisation. Financial support for the work was obtained from the National Natural Science Foundation of China (NSFC) grant (30700664) and the National High-tech Research and Development Project (2007AA06Z404) of China.