Production of Sensitive Monoclonal Antibodies to Aflatoxin B1and Aflatoxin M1 and Their Application to ELISA of Naturally Contaminated Foods

Abstract

Two new hybridoma Cell lines capable of secreting sensitive monoclonal antibodies for aflatoxin B1 (AFB1) and aflatoxin M1 (AFM1), were produced by fusing NS-1 myeloma cells with spleen cells of BALB/c female mice immunized with AFB1- and AFM1-carboxymethyloxime bovine serum albumin conjugates, respectively. Detection limits for these antibodies in the direct enzyme-linked immunosorbent assay (ELISA) were 0.5 ng/ml for AFB1 and 0.25 ng/ml for AFM1 Concentrations of AFB1 analogs (ng/ml) required to inhibit 50% binding of AFB1,-perioxidase conjugate to AFB1 monoclonal antibody solid phase in direct ELISA were: AFB1, 2.6; AFB2, 13; AFG1, 8; AFB2, 15; AFM1, 23. Analog concentrations (ng/ml) required to inhibit 50% binding of AFB1,-perioxidase conjugate to AFM1 monoclonal antibody solid phase were: AFM1,0.8; AFM2, 700; AFB1, 0.5; AFB2, 35; AFB2a, >10,000; AFG1, 12; AFG2a, 12; AFP1, 16; and AFQ1, 9.2. These new monoclonal antibodies were applicable to both the ELISA detection AFB1 in corn, cottonseed, cottonseed meal, and mixed feed following a simple extraction in 55% methanol as well as the direct detection of AFM1 in milk.