A wash-free and label-free colorimetric biosensor for naked-eye detection of aflatoxin B1 using G-quadruplex as the signal reporter

•This is a wash-free and label-free biosensor with simple operation.•The results can be directly visualized by the naked eye.•The assay is ultrasensitive, with a detection limit of 1 pM.•This system is robust and can be applied to the determination of AFB1 in peanut samples. A wash-free and label-free colorimetric biosensor for the amplified detection of aflatoxin B1 (AFB1) has been constructed by the integration of an ingenious hairpin DNA probe with exonuclease III (Exo III)-assisted signal amplification. The presence of the AFB1 activates the continuous cleavage reactions by Exo III toward a hairpin probe, resulting in the autonomous accumulation of numerous free G-quadruplex sequences, which can catalyze the oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB) by H2O2 to produce a colorimetric response. The naked-eye biosensor is ultrasensitive, enabling the visual detection of trace amounts of AFB1 as low as 1 pM without instrumentation. The sensor is robust and can work even when challenged with complex sample matrices such as peanut samples. With the advantages of simple operation, wash-free and label-free format, visible and intuitive output, and low cost, the naked-eye based colorimetric biosensor is expected to have potential applications for in-field detection of AFB1.