Development and application of an enzyme-linked immunosorbent assay for the analysis of hydrolyzed fumonisins

Abstract

Polyclonal antibodies against hydrolyzed fumonisin B₁ (HFB₁) were prepared by immunization of rabbits with a keyhole limpet hemocyanin conjugate coupled by glutaraldehyde. Sensitivity and specificity of these antibodies were tested in a direct competitive enzyme-linked immunosorbent assay (ELISA), in which a HFB₁-horseradish peroxidase conjugate prepared by reductive alkylation served as the labeled antigen. A direct competitive ELISA for the quantitative determination of HFB₁ in corn-based food samples was developed with this antibody. The 50% inhibition level was used for the determination of cross-reactivity. The cross-reactivities of the antibodies for HFB₂ and HFB₃ were 4.3% and 14.4%, respectively, whereas the parent compound fumonisin B₁ (FB₁) showed no cross reactivity. The detection limit of the EIA calculated from the HFB₁-concentration given a 30% binding inhibition was 0.25 ng/ml for the standard solutions and 10 ng/g for the analyzed food samples e.g. tortilla chips, nachos and cornflakes. Mean recoveries of HFB₁ for artificially contaminated food samples were in the range of 96 to 98%. This test offers a rapid and economic opportunity for the HFB₁-screening of food samples.