Preparation for aflatoxin B1-cationized bovine serum albumin based on Mannich-type reaction

Abstract Aflatoxin B 1 (AFB 1 ), which is commonly found in agricultural commodities, is one of the most potent carcinogenic mycotoxins. To ensure food safety, rapid and low-cost immunological methods have been applied to detect AFB 1 worldwide. A key step in these immunological methods is coupling AFB 1 to carrier proteins; AFB 1 is usually deviated to AFB 1 -oxime and coupled to carrier proteins to form the AFB 1 -oxime–protein conjugate. In the current research, AFB 1 was directly coupled with cationized bovine serum albumin (cBSA) using a method based on Mannich-type principles. The coupling effects were investigated with different initial molar ratios of AFB 1 to cBSA. The conjugate molar ratio was 6.4:1 when the initial molar ratio was 40:1. The cationized proteins and their conjugates were identified by UV–Vis and FT-IR spectra, which showed the characteristic bands of the ethylendiamine group and AFB 1 , respectively. After BALB/c mice were immunized with AFB 1 –cBSA, a quicker immunological response and a similar sensitivity of antisera against AFB 1 were observed, compared with immunization by AFB 1 -oxime–BSA. This suggests that the Mannich-type reaction might be an alternative method of preparation for AFB 1 –protein.