Sensitive competitive direct enzyme-linked immunosorbent assay and gold nanoparticle immunochromatographic strip for detecting aflatoxin M1 in milk

Polyclonal antibodies specific to aflatoxin M1 were generated from rabbits immunized with AFM1-bovine serum albumin (BSA). By using these antibodies, this work establishes a rapid competitive direct enzyme-linked immunosorbent assay (cdELISA) and a gold nanoparticle immunochromatographic strip method for detecting AFM1 in milk and milk products. In the rapid cdELISA, AFM1 at a concentration of 0.014 ng/ml caused 50% inhibition (IC50) of binding AFM1-horseradish peroxidase to the antibodies. Effective on-site detection of AFM1 was also developed based on a rapid and sensitive antibody-gold nanoparticle immunochromatographic strip method. This strip had a detection limit of 1.0 ng/ml for AFM1 in milk samples. Additionally, the whole analysis was completed within 10 min. Close examining 15 milk-based samples by cdELISA revealed that 6 were slightly contaminated with AFM1, with a mean concentration of 0.028 ng/ml. Moreover, all samples were negative in the immunochromatographic strip assays due to the contaminated levels were below the detection limit of the strip. Importantly, the proposed cdELISA and immunochromatographic strip methods are highly sensitive to the rapid detection of AFM1 in milk and milk products.