An enzyme-linked immunosorbent assay and a gold-nanoparticle based immunochromatographic test for amatoxins using recombinant antibody

The authors describe two kinds of rapid assays for the determination of amatoxins in mushrooms. The first is an enzyme-linked immunosorbent assay (ELISA) using horseradish peroxidase. The second is a rapid immunochromatographic assay that uses colloidal gold as a red label (CG-ICA). Both are based on the use of a well-characterized recombinant single chain variable fragment antibody (named scFv-A4). The half-maximum inhibition concentrations (IC50) of alpha-amanitin, beta-amanitin and gamma-amanitin are 78, 85 and 90 ng.mL(-1), and the limits of detection (LODs; for IC15) are 1.9, 2.1 and 2.8 ng.mL(-1). The method was applied to the determination of amanitins in mushrooms, and the LODs for alpha-amanitin, beta-amanitin and gamma-amanitin in mushroom samples were found to be 4.9, 6.4 and 8.3 ng.mL(-1). The visual minimum detection limits of the optimized CGIA are 4 and 6 ng.mL(-1) for mushroom samples. The test can be performed within 10 min. The results of the analysis of spiked samples showed that the CG-IA can rapidly and semi-quantitatively quantify amatoxins in mushroom samples on site and at low costs.