Engineering DNA G-quadruplex assembly for label-free detection of Ochratoxin A in colorimetric and fluorescent dual modes

Colorimetric and fluorescent methods for Ochratoxin A (OTA) detection are convenient and well received. However, the pigments and autofluorescence originated from food matrices often interfere with detection signals. We have developed a strategy with colorimetric and fluorescent dual modes to solve this challenge. In the colorimetric mode, OTA aptamer (AP9) was assembled into a DNA triple-helix switch with a specially designed signal-amplifying sequence. The OTA-induced G-quadruplex (G4) of AP9 would open the switch and release the signal-amplifying sequence for colorimetric signal amplification. The G4 structures of AP9 were further utilized to combine with the fluorogenic dye ThT for fluorescent mode. By skillfully engineering DNA G4 assembly for signal amplification, there was no need for any DNA amplification or nanomaterials labeling. Detections could be carried out in a wide temperature range (22-37 ℃) and finished rapidly (colorimetric mode, 60 min; fluorescent mode, 15 min). Broad linear ranges (colorimetric mode, 10-1.5 ×103 μg/kg; fluorescent mode, 0.05-1.0 ×103 μg/kg) were achieved. The limit of detection for colorimetric and fluorescent modes were 4 μg/kg and 0.01 μg/kg, respectively. The two modes have been successfully applied to detect OTA in samples with intrinsic pigments and autofluorescence, showing their applicability and reliability.