CRISPR/Cas12a and G-quadruplex DNAzyme-driven multimodal biosensor for visual detection of Aflatoxin B1.

Aflatoxin B1 (AFB1) contamination severely threatens human and animal health, it is thus critical to construct a strategy for its rapid, accurate, and visual detection. Herein, a multimodal biosensor was proposed based on CRISPR/Cas12a cleaved G-quadruplex (G4) for AFB1 detection. Briefly, specific binding of AFB1 to the aptamer occupied the binding site of the complementary DNA (cDNA), and cDNA then activated Cas12a to cleave G4 into fragments. Meanwhile, the intact G4-DNAzyme could catalyze 3, 3', 5, 5'-tetramethylbenzidine (TMB) to form colourimetric/SERS/fluorescent signal-enhanced TMBox, and the yellow solution produced by TMBox under acidic conditions could be integrated with a smartphone application for visual detection. The colourimetric/SERS/fluorescent biosensor yielded detection limits of 0.85, 0.79, and 1.65 pg·mL-1, respectively, and was applied for detecting AFB1 in peanut, maize, and badam samples. The method is suitable for visual detection in naturally contaminated peanut samples and has prospective applications in the food industry.