An Indirect Enzyme-Linked Immunosorbent Assay for T-2 Toxin in Biological Fluids

Abstract

An indirect enzyme-linked immunosorbent assay (ELISA) which can detect 0.2 to 1 ng of T-2 toxin per ml in urine, serum and milk was developed. T-2 hemisuccinate was conjugated to polylysine which was then coated to a microtiter plate and incubated with rabbit anti-T-2 antibody and sample extract. The amount of anti-T-2 antibody bound to the plate was then determined by reaction with goat anti-rabbit IgG-peroxidase complex and by subsequent reaction with the substrate. Samples spiked with T-2 toxin were subjected to a simple cleanup procedure by passing them through a reversed-phase Sep-Pak catridge (C18). The recoveries of tritiated T-2 toxin added to the urine, serum and milk samples were between 71 to 90% after the cleanup step. In the ELISA, significant interference was observed when more than 5μl of sample, without cleanup treatment, were used in each analysis. After cleanup, extracts equivalent to 50μl of serum, urine or milk per well did not significantly interfere with the assay. The recoveries of T-2 toxin added to serum (1 to 10 ng/ml), urine (0.2 to 10 ng/ml) and milk (0.2 to 10 ng/ml) after cleanup treatment as determined by the indirect ELISA were found to be 51 to 82%, 73 to 82% and 80 to 83%, respectively.