A DNA-Tile-Based Fluorescence Lighting System for Signal-Amplified Imaging at the Single Live-Cell Level

Nucleic acid aptamer technology, with precise targeting and excellent biosafety, has significantly improved dynamic fluorescence imaging at the live-cell level. While integrating isothermal amplification technology can effectively enhance target signal output to improve imaging effects, the high sequence homology with aptamers often increases the background signal and presents a significant challenge for sequence design. Herein, we developed a novel DNA tile-based fluorescent labeling system (DTFL system), in which the targeting module carrying aptamers recognizes and binds specific cellular markers and the signal-amplified module modified with fluorescent groups acts as fuel to rapidly initiate melamine-responsive array assembly on the membrane surface, realizing nanonet-encapsulated imaging on single living cells. This system demonstrated significant superiority in rapid cellular imaging, attributable to melamine-mediated T-base mismatches, forming bonds more readily than classical Watson-Crick base pairing. Besides, the separation of targeting and imaging elements ensures sufficient signal intensity while avoiding the cellular internalization of fluorescence signals, ultimately improving the imaging sensitivity and accuracy. Under optimal reaction parameters, this system was applied to real-time imaging of various kinds of living cells and served as a dynamic monitoring tool to precisely track changes in the expression levels of specific markers. With a simple sequence design, easy operation, and excellent biological safety, the system can be readily implemented in fundamental laboratory settings and represents a highly promising imaging strategy to enrich the toolbox for live-cell analysis.