Development of an Electrochemical Biosensor for the Detection of Aflatoxin M1 in Milk

Abstract

We have developed an electrochemical immunosensor for the detection of ultratrace amounts of aflatoxin M1 (AFM1) in food products. The sensor was based on a competitive immunoassay using horseradish peroxidase (HRP) as a tag. Magnetic nanoparticles coated with antibody (anti-AFM1) were used to separate the bound and unbound fractions. The samples containing AFM1 were incubated with a fixed amount of antibody and tracer [AFM1 linked to HRP (conjugate)] until the system reached equilibrium. Competition occurs between the antigen (AFM1) 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 AFM1-contaminated milk from the ELISA kit was used to obtain a standard curve for AFM1. 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 AFM1. Our immunosensor has a low detection limit (0.01 ppb), which is under the recommended level of AFM1 [0.05 µg L-1 (ppb)], and has good reproducibility.

Keywords

• electrochemical immunosensor
• aflatoxin M1
• mycotoxin
• milk
• horseradish peroxidase (HRP)
• superparamagnetic nanoparticles