Self-assembled DNA origami-based duplexed aptasensors combined with centrifugal filters for efficient and rechargeable ATP detection

DNA origami technology has great potential for biosensor applications. Here, we described the construction of a self-assembled DNA origami biosensor for the precise localization of fluorescent aptamers. Due to the molecular weight difference between DNA origami and aptamer, centrifugal filters were used to quantitatively detect adenosine triphosphate (ATP). The ATP-specific aptamer labeled with fluorescence reporter 6-carboxyfluorescein FAM (FAM-aptamer) was selected as the recognition element and signal probe. ATP duplexed aptamers bound to triangular DNA origami by base-complementary pairing, resulting in high fluorescence signals on the origami arrays. The competitive binding of ATP toward the FAM-aptamer triggered the release of FAM-aptamer-ATP complexes from the surface of the origami array, resulting in weakened fluorescence signals. For ATP quantification, 100 kD centrifugal filters were employed, followed by measurement of the fluorescence signal trapped on the origami arrays of the filter device. The successful synthesis of origami-aptamer arrays was characterized by atomic force microscopy, laser confocal microscopy, and electrophoresis. Fluorescence measurements exhibited an excellent linear relationship with logarithms of ATP concentrations within 0.1-100 ng mL-1, with a detection limit of 0.29 ng mL-1. By replacing aptamers and complementary strands, we demonstrated the potential of this method for 17β-estradiol detection. Considering that the detection mechanism is based on the hybridization and displacement of DNA strands, the detection system had the potential for recharging. Our study provides new insights into applying DNA origami technology in small molecule detection.