Development of a simple and convenient cell-based electrochemical biosensor for evaluating the individual and combined toxicity of DON, ZEN, and AFB1

A simple and convenient cell-based electrochemical biosensor was developed to assess the individual and combined toxicity of deoxynivalenol (DON), zearalenone (ZEN), and Afiatoxin B-1 (AFB(1)) on Hep G2 cells. The sensor was modified in succession with AuNPs (gold nanoparticles), cysteamine, and laminin. The cells interacting with laminin formed tight cell-to-electrode contacts, and collagen was used to maintain cell adhesion and viability. Electrochemical impedance spectroscopy (EIS) was developed to evaluate mycotoxin toxicity. Experimental results show that DON, ZEN, and AFB(1) caused a significant decrease in cell viability in a dose dependent manner. The EIS value decreased with concentrations of DON, ZEN, and AFB(1) in the range of 0.01-20, 0.1-50, and 0.1-3.5 mu g/mL, and IC50 obtained using the developed method was 48.5, 59.0, and 3.10 mu g/mL, respectively. A synergistic effect was observed between DON and ZEN, an additive effect was observed between DON and AFB(1), and an antagonism effect was found in the binary mixtures of ZEN and AFB(1) and ternary mixtures. These results were confirmed via CCK-8 assay. Utilizing SEM, we found that cells treated with mycotoxins caused significant changes in cell morphology, thus lessening cell adsorption and impedance reduction. Biological assay indicated that EIS patterns correlated with [Ca2+](i) concentrations and apoptosis and necrotic cells ratios, thus effecting electrochemical signals. This method is simpler, more convenient, sensitive, and has a quicker response rate than most conventional cytotoxicity evaluation methods.