Development of an Immunofluorescence Lateral Flow Assay for Detecting Four Medically Important Snake Venoms in China

Snakebite envenomation requires rapid venom identification for effective treatment, a need unmet by current diagnostics in China. This study aimed to develop a rapid, multiplex immunofluorescence lateral flow assay for the simultaneous detection of four major venomous snakes: Deinagkistrodon acutus, Bungarus multicinctus, Naja atra , and Agkistrodon halys . Key findings demonstrated that the assays exhibited high sensitivity, with limits of detection (LODs) ranging from 0.6 to 1.2 ng/mL. Excellent specificity was confirmed, with no cross‐reactivity observed among the four venoms. In a rabbit envenomation model, the assays accurately quantified venom in various biological samples (venous serum, wound exudate) within 5 min. Receiver operating characteristic (ROC) curve analysis showed high diagnostic accuracy, with areas under the ROC curve (AUCs) ranging from 0.889 to 0.956. Assay accuracy was further verified by spike‐in recovery rates of 90.3%–109.6%. The results indicate that this novel multiplex immunoassay shows great potential as a point‐of‐care tool for the rapid and quantitative diagnosis of snakebites, showing great potential for preclinical application following further human sample validation.