Detection of Aflatoxin B1 in Corn, Rice, and Barley by ELISA, Using a Heavy-Chain IgG2b Isotype Monoclonal Antibody

Abstract

A rapid and sensitive, indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) method has been developed to measure aflatoxin B1 (AFB1) in corn, rice, and barley using a heavy-chain IgG2b isotype monoclonal antibody (MAb). One MAb was isolated and characterized after fusion of myeloma cells with spleen cells isolated from BALB/c mice that had been immunized with AFB1 carboxymethyl oxime conjugated with bovine serum albumin. The antibody was IgG2b with a λ light chain. The crossreactivities of the anti-AFB1 MAb clone were 1.0% against aflatoxin M1 and <1.0% against aflatoxin B2, aflatoxin G1, and deoxynivalenol. The limit of detection for AFB1 was 0.001 ng/mL, and the AFB1 concentration required for 50% inhibition of binding was 0.01 ng/mL. The linear range for the developed IC-ELISA was 0.001–10.0 ng of AFB1 per mL. Assays of cereal extract samples mixed with AFB1 ranging in concentration from 0.1 to 10 ng/mL resulted in a mean ELISA recovery of 93.7%. In total, 53 samples of corn, rice, and barley were collected from farms in northeastern China. The concentration of AFB1 in all samples collected was <1 ng/g. The average AFB1 contamination rates were 66% in corn, 42% in rice, and 58% in barley. The results indicate that necessary precautions will have to be taken to minimize AFB1 contamination in corn, rice, and barley.