Production of highly sensitive monoclonal antibody and development of lateral flow assays for phallotoxin detection in urine.

Abstract

Phallotoxins, toxic cyclopeptides found in wild poisonous mushrooms, are predominant causes of fatal food poisoning. For the early and rapid diagnosis mushroom toxin poisoning, a highly sensitive and robust monoclonal antibody (mAb) against phallotoxins was produced for the first time. The half-maximum inhibition concentration (IC50) values of the mAb-based indirect competitive ELISAs for phallacidin (PCD) and phalloidin (PHD) detection were 0.31ngmL-1 and 0.35ngmL-1, respectively. In response to the demand for rapid screening of the type of poisoning and accurate determination of the severity of poisoning, colloidal gold nanoparticle (GNP) and time-resolved fluorescent nanosphere (TRFN) based lateral flow assays (LFA) were developed. The GNP-LFA has a visual cut-off value of 3.0ngmL-1 for phallotoxins in human urine sample. The TRFN-LFA provides a quantitative readout signal with detection limit of 0.1ngmL-1 in human urine sample. In this study, urine samples without pretreatment were used directly for the LFA strip tests, and both two LFAs were able to accomplish analysis within 10min. The results demonstrated that LFAs based on the newly produced, highly sensitive, and robust mAb were able to be used for both rapid qualitative screening of the type of poisoning and accurate quantitative determination of the severity of poisoning after accidental ingestion by patients of toxic mushrooms.