Development of ELISA and colloidal gold immunoassay for tetrodotoxin detetcion based on monoclonal antibody

A monoclonal hybridoma cell named 5B9 against tetrodotoxin (TTX) was obtained after fusion of myeloma SP2/0 cells with spleen cells isolated from the immunized Balb/c mice. The 5B9 monoclonal antibody (McAb) with high affinity (about 2.55×109) is specific to TTX, and this McAb belongs to the immunoglobulin G (IgG) isotype. Finally, an enzyme-linked immunosorbent assay (ELISA) and colloidal gold immunoassay were established based on this McAb. The linear range of ELISA to detect TTX was 5-500 ng/mL, and the limit of detection (LOD) was 4.44 ng/mL. The average CV of intra- and inter-assay was less than 8%, with the samples recovery range of 70.93-99.99%. A competitive format colloidal gold strip was developed for detection of TTX in real samples, and the LOD for TTX is 20 ng/mL, and the assay time of the qualitative test can be finished in less than 10 min without any equipment. The result from test strip revealed that the test strip has a good agreement with those obtained from ELISA.