Quantitative Super‐Resolution Imaging of On‐Origami DNA Conformation and Reactivity Under Electric Fields

Abstract Self‐assembled DNA nanostructures have been popularly used to develop DNA‐based electrochemical sensors by exploiting the nanoscale positioning capability of DNA origami. However, the impact of the electric field on the structural stability of the DNA origami framework and the activity of carried DNA probes remains to be explored. Herein, we employ DNA origami as structural frameworks for reversible DNA hybridization, and develop a single‐molecule fluorescence imaging method to quantify electric field effects on DNA conformation and hybridization properties at the single‐molecule level. Through single‐molecule temporal kinetic analysis of hybridization events occurring on individual DNA origami, we systematically determine the regulation patterns of applied potential and scanning duration on the activity of DNA probes. Optical super‐resolution reconstruction of probe sites reveals electric field‐induced structural relaxation in DNA frameworks. This approach not only provides insights into electrochemical DNA sensing devices, but also lays the foundation for developing hybrid electrical–optical analysis at the single‐molecule level.