Construction of a fluorescent and colorimetric multi-mode aptasensor for label-free and sensitive detection of ochratoxin A in agricultural products

Ochratoxin A (OTA) can induce a serious danger to human beings, and it is crucial to develop a sensitive and simple strategy for visual detection of OTA. Herein, we construct a fluorescent and colorimetric multi-mode aptasensor for label-free and sensitive detection of OTA on the basis of target-initiated assembly of functional G-quadruplex (G4) DNAzyme nanostructure. When OTA is present, it specifically recognizes the aptamer and disintegrates the aptamer-cDNA duplex structure, resulting in the liberation of intact cDNA and the subsequent initiation of terminal deoxynucleotidyl transferase (TdT) assist-extension reaction. The resultant G4 DNAzyme nanostructure can light up Thioflavin T (ThT) to produce a high fluorescence signal. Meanwhile, the addition of hemin can trigger the self-assembly of hemin/G4 DNAzymes that can catalyze the 3,3′,5,5′-tetramethylbenzidine/hydrogen peroxide (TMB/H2O2) system to generate a distinct colorimetric signal. Consequently, the introduction of G4 DNAzyme/ThT and hemin/G4 DNAzyme can efficiently transduce the OTA signal into both a fluorescence signal and a colorimetric signal. The further utilization of a smartphone facilitates the development of a simple method based on red-green-blue (RGB) digital analysis for OTA assay. The proposed multi-mode aptasensor displays ultrahigh sensitivity with a detection limit of 2 pg/mL for the fluorescent mode, 25 pg/mL for the colorimetric mode, and 6 pg/mL for the RGB mode. Moreover, this multi-mode aptasensor can be applied to accurately detect OTA in real samples, providing a new platform for on-site monitoring of OTA in agricultural products.