One of the most poisonous mushrooms is the death cap mushroom (<i>Amanita phalloides).</i> The most toxic constituent is α-amanitin, a bicyclic octapeptide, which damages the liver and kidneys. To develop a new tool for detecting this toxin, monoclonal antibodies (mAbs) were developed and characterized. Both α- and b-amanitin were coupled to carrier proteins through five different linking chemistries. These conjugates were evaluated for their effectiveness in generating antibodies specific for the free toxin, as well as their utility in formatting heterogeneous assays. Ultimately, these efforts yielded a newly described conjugation procedure that successfully resulted in generating sensitive mAbs. Analysis of the conjugates by MALDI-TOF confirmed the linkage and estimated the hapten loading to be approximately 4 haptens per BSA. In addition, the mAbs were characterized for their sensitivity and selectivity using the mAbs in a competitive ELISA format. These experiments revealed that α-amanitin was detected at sub-ppb (ng/mL) concentrations.

Development of an immunoassay for amanitin, a highly toxic mushroom toxin

255th National Meeting (2018)

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