[Using HTS-ELISA method to make anti-aflatoxin M1 monoclonal antibody].

OBJECTIVE: To prepare high-affinity anti-aflatoxin M1 monoclonal antibodies by High Throughput Screening ELISA (HTS-ELISA) METHODS: Balb/C mice were immunized by aflatoxin M1-bovine serum albumin conjugate, and screen secret anti-aflatoxin M1 monoclonal antibody hybridoma by HTS-ELISA. The antibody was characterized. RESULTS: Fourteen hybridoma cell lines which could secret high activity anti-aflatoxin M1 monoclonal antibodies were obtained. The affinity of the purified monoclonal antibody was 5.5 x 10(-10) mol/L. The cross-reactivity of the monoclonal antibody clone against aflatoxin M1, aflatoxin M2, aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2, deoxynivalenol and BSA was 100%, 4.5%, 21.5%, 1.0%, 16.6%, 1.0%, 0%, 0%, respectively. The sensitivity of the anti-AFM1 monoclonal antibody binding to aflatoxin M1 was 0.01 microg/L and the linear range for developed indirect competitive ELISA was 0.1 - 10 microg/L aflatoxin M1. The binding inhibition IC50 of the anti-aflatoxin M1 monoclonal antibody was 0.82 microg/L. Assays of milk samples mixed with AFM1 ranging in concentration from 0.25 to 5.0 microg/L gave mean indirect competitive ELISA recovery of 60.3% - 152.8%. CONCLUSION: HTS-ELISA can be used for the preparation of the high-affinity anti-aflatoxin M1 monoclonal antibodies. The anti-aflatoxin M1 monoclonal antibody could be provided as the high quality material in the system of aflatoxin M1 immune detection.