Abstract: A rapid and sensitive direct competitive enzyme-linked immunosorbent assay (ELISA) using anti-AFM1 mAb with horseradish peroxidase (HRP) to measure aflatoxin M1 (AFM1) in milk was described. Using AFM1 contaminated milk ERMI-BD282 (zero), ERMI-BD 283 (0.11 μg/kg) and ERMI-BD 284 (0.44μg/kg), the sensitivity and accuracy of the developed assay was validated. The optimized assay conditions regarding sensitivity and stability were found to be: coating AFM1-BSA antigen concentration 0.25 μg/kg and dilution factor of anti-AFM1 mAb-HRP conjugate 2000. Assays of ERMI-BD282 samples spiked with AFM1 at the level of 0.45 μg/L revealed an average recovery of around 80%. The developed method showed an IC50 of 0.75μg/L and a linear range of 0.015－4.05 μg/L. This assay may be used in rapid screening of contaminated milk at AFM1 ＞0.5μg/L.

Key words: direct competitive enzyme-linked immunosorbent assay (dc-ELISA), aflatoxin M1 (AFM1), milk, detection