Development of an Electrochemical Biosensor for the Detection of Aflatoxin M ₁ in Milk

Abstract

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.