Submitted to: Toxins
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/9/2019
Publication Date: 12/11/2019
Citation: Bever, C.R., Hnasko, R.M., Cheng, L.W., Stanker, L.H. 2019. A rapid extraction method combined with a monoclonal antibody-based immunoassay for the detection of amatoxins. Toxins. 11(12). https://doi.org/10.3390/toxins11120724.
Interpretive Summary: A variety of mushrooms contain deadly toxins, known as amatoxins. Methods to quickly detect these toxins are needed for food safety. Here we report the development of a monoclonal antibody-based assay with a detection limit in the ppb (ng/mL) range. Rapid and simple sample preparation methods were developed and tested. Toxins from small pieces of mushrooms were extracted using solutions as simple as just water, shaken by hand for only a minute. Together, the extraction method and the monoclonal antibody assay represents a simple and fast test that readily detects amatoxins extracted from mushroom samples.
Technical Abstract: Amatoxins are lethal toxins found in a variety of mushroom species. Rapid and sensitive methods to detect amatoxins are needed to quickly determine if amatoxins are present in cases of suspected mushroom intoxications. Antibody-based assays permit rapid and sensitive detection. In this work, we demonstrate the production of new monoclonal antibodies that are used in a competitive enzyme-linked immunosorbent assay (cELISA) that is sensitive at 1 ppb (1 ng/mL) and shows selectivity for a-amanitin (a-AMA), ß-amanitin (ß-AMA) and '-amanitin ('-AMA). In order to decrease the overall time needed for analysis, the extraction procedure for mushrooms also was simplified. A rapid (a few minutes) extraction of amatoxins from mushroom samples using buffers - as simple as only water - were developed and was successful for detecting toxins from Amanita phalloides and A. ocreata. The extracts containing toxins required dilution of at least 10,000-fold before a change in signal intensity was observed in the cELISA. Together, the extraction method and the monoclonal antibody ELISA represents a simple and fast test that readily detects amatoxins extracted from mushroom samples.