Development of Surface Plasmon Resonance-Based Immunoassay for Aflatoxin B1

Abstract

Aflatoxins are a group of highly toxic fungal secondary metabolites that occur in Aspergillus species and may contaminate foodstuffs and feeds. Two different anti-aflatoxin B₁ antibodies were examined to develop a surface plasmon resonance (SPR)-based immunoassay to aflatoxin B₁. A conjugate consisting of aflatoxin B₁−bovine serum albumin (BSA) was immobilized on the dextran gel surface. Competition between immobilized aflatoxin B₁ conjugate and free aflatoxin B₁ 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 NaOH and HCl worked to a degree, but regeneration was at the expense of the integrity of the immobilized conjugate. A polyclonal anti-aflatoxin B₁ antibody 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 combined high ionic strength and extreme pH, as well as chaotrophic properties and allowed the development of an inhibitive immunoassay. The assay had a linear range of 3.0−98.0 ng mL^-1 with good reproducibility. Keywords: Aflatoxin B₁; surface plasmon resonance; regeneration; inhibitive immunoassay