Development of Monoclonal Antibody-based Enzyme-linked Immunosorbent Assay for Detecting Tetrodotoxin

This study aimed to develop a method for the detection of tetrodotoxin (TTX) based on monoclonal antibody (McAb). The hapten TTX was linked to carrier protein keyhole limpet hemocyanin (KLH) as immunogen, and linked to ovalbumin (OVA) as coating antigen by the Mannich method. Then the 6 similar to 8 weeks Babl/c mice were immunized. After cell amalgamation, a cell line with high specificity and sensitivity was obtained, and McAb was produced. The indirect competitive enzyme-linked immunosorbent assay (icELISA) was developed for detecting TTX. The optimized working conditions of the icELISA were 4.0 mu g/mL coating antigen, 0.75 mu g/ML McAb, 45 min competitive time, at room temperature (20 similar to 25 degrees C). The IC50 value of this method was 24.0 ng/mL, the working ranges were 5.2 similar to 107.6 ng/mL, the intra-assay and inter-assay coefficient of variation (CV %) were 4.2 and 4.5, respectively. This investigation will benefit the assay kit development for detecting TTX.