We have developed an electrochemical immunosensor for the detection of ultratrace amounts of aflatoxin M(1) (AFM(1)) in food products. The sensor was based on a competitive immunoassay using horseradish peroxidase (HRP) as a tag. Magnetic nanoparticles coated with antibody (anti-AFM(1)) were used to separate the bound and unbound fractions. The samples containing AFM(1) were incubated with a fixed amount of antibody and tracer [AFM(1) linked to HRP (conjugate)] until the system reached equilibrium. Competition occurs between the antigen (AFM(1)) and the conjugate for the antibody. Then, the mixture was deposited on the surface of screen-printed carbon electrodes, and the mediator [5-methylphenazinium methyl sulphate (MPMS)] was added. The enzymatic response was measured amperometrically. A standard range (0, 0.005, 0.01, 0.025, 0.05, 0.1, 0.25, 0.3, 0.4 and 0.5 ppb) of AFM(1)-contaminated milk from the ELISA kit was used to obtain a standard curve for AFM(1). To test the detection sensitivity of our sensor, samples of commercial milk were supplemented at 0.01, 0.025, 0.05 or 0.1 ppb with AFM(1). Our immunosensor has a low detection limit (0.01 ppb), which is under the recommended level of AFM(1) [0.05 μg L-1 (ppb)], and has good reproducibility.
Keywords: aflatoxin M1; electrochemical immunosensor; horseradish peroxidase (HRP); milk; mycotoxin; superparamagnetic nanoparticles.