Poly-thymine-mediated fluorescence recovery of N-CQDs for turn-on sensing of APE1 activity in breast cancer cell lysates

A label-free turn-on fluorescent biosensor is developed for APE1 activity detection based on APE1-gated release of poly-T strands from a constrained DNA duplex, which competitively displaces Ag⁺ from N-CQDs, recovering fluorescence. The sensor achieves a detection limit of 0.02 U mL⁻¹, excellent selectivity, and one-step operation. It successfully detects APE1 in serum (recoveries 92-108%) and in cancer cell lysates (MCF-7, MDA-MB-231, HeLa, HepG2), with a threshold as low as 192 cells, and distinguishes cancer cells from normal cells. This simple, robust platform holds promise for DNA repair research, cancer diagnostics, and inhibitor screening.