Development of surface plasmon resonance-based immunoassay for aflatoxin B-1

Aflatoxins are a group of highly toxic fungal secondary metabolites that oc cur in Aspergillus species and may contaminate foodstuffs and feeds. Two di fferent anti-aflatoxin B-1 antibodies were examined to develop a surface pl asmon resonance (SPR)-based immunoassay to aflatoxin B-1. A conjugate consi sting of aflatoxin Br-bovine serum albumin (BSA) was immobilized on the dex tran gel surface. Competition between immobilized aflatoxin B-1 conjugate a nd free aflatoxin B-1 in solution for binding to antibody injected over the surface formed the basis for the assay. Regeneration of the antibody from the immobilized conjugate surface is essential for the development of such an inhibitive immunoassay. Problems were encountered with the regeneration of the sensor surface, due to the high-affinity binding of the antibodies. Conventional regeneration solutions consisting of low concentrations of NaO H and HCl worked to a degree, but regeneration was at the expense of the in tegrity of the immobilized conjugate. A polyclonal anti-aflatoxin B-1 antib ody was produced and was found to be regenerable using an organic solution consisting of 1 M ethanolamine with 20% (v/v) acetonitrile, pH 12.0. This c ombined high ionic strength and extreme pH, as well as chaotrophic properti es and allowed the development of an inhibitive immunoassay. The assay had a Linear range of 3.0-98.0 ng mL(-1) with goad reproducibility.