Visual and semiquantitative spectrophotometric ELISA screening method for aflatoxin B1 in corn and peanut products: follow-up collaborative study.

Abstract

A joint AOAC/IUPAC (International Union of Pure and Applied Chemistry) interlaboratory study of an enzyme-linked immunosorbent screening assay (ELISA) for aflatoxins was conducted in laboratories in Canada, France, Japan, The Netherlands, Switzerland, Tunisia, and the United States. Twelve raw and roasted peanut and corn portions containing various concentrations of natural aflatoxins and supplemented when appropriate with aflatoxin B1 were distributed to participating laboratories for testing. The assay is based on competition between an enzyme-conjugated aflatoxin B1 and (free) aflatoxins in the test sample for aflatoxin-specific antibodies coated onto interior surfaces of microtiter wells. After a wash step to remove all unbound aflatoxins, a substrate added to each well is catalyzed from a colorless to a blue solution by any bound enzyme-conjugated aflatoxin B1 present. The intensity of the color decreases as the amount of free aflatoxin B1 in the test portion increases. Final determination of aflatoxin concentrations can be made by either visual comparison with standard solutions or spectrophotometric comparisons (at 650 nm) to knowns. Overall correlation was good between ELISA and thin-layer chromatographic results for corn and roasted peanut products, with 93 and 98% correct responses for visual and instrumental determinations, respectively. For instrumental determinations of aflatoxin in corn and roasted peanuts in the less than or equal to 20 ng/g range, the relative standard deviations for repeatability (RSDr) were 14.9 and 41.4%, respectively, and the relative standard deviations for reproducibility (RSDR) were 45.7 and 43.5%, respectively. For instrumental determination of greater than 20 ng/g, the respective RSDr and RSDR values were 19.4 and 52.7% for corn and 23.3 and 23.3% for roasted peanuts.(ABSTRACT TRUNCATED AT 250 WORDS)