Nitroso-Caged Rhodamine: A Superior Green Light-Activatable Fluorophore for Single-Molecule Localization...

The evolution of super-resolution imaging techniques, especially single-molecule localization microscopy, demands the engineering of switchable fluorophores with labeling functionality. Yet, the switching of these fluorophores depends on the exterior conditions of UV light and enhancing buffers, which is bioincompatible for living-cell applications. Herein, to surpass these limitations, a nitroso-caging strategy is employed to cage rhodamines into leuco forms, which for the first time, is discovered to uncage highly bright zwitterions by green light. Further, clickable construction grants the specificity and versatility for labeling various components in living cells. The simultaneous photoactivation and excitation of these novel probes allow for single-laser super-resolution imaging without any harmful additives. Super-resolution imaging of microtubules in fixed cells or mitochondria and the distribution of glycans and H2B proteins in living cells are achieved at a molecular scale with robust integrity. We envision that our nitroso-caging probes would set a platform for the development of future visible-activatable probes.