No-wash fluorogenic labeling of proteins for reversible photoswitching in live cells

Photoswitchable fluorescent molecules (PSFMs) are positioned as valuable tools for biomolecule localization tracking and super-resolution imaging technologies due to their unique ability to reversibly control fluorescence intensity upon light irradiation. Despite the high demand for PSFMs that are suitable for live-cell imaging, no general method has been reported that enables reversible fluorescence control on proteins of interest in living cells. Herein, we have established a platform to realize reversible fluorescence switching in living cells by adapting a protein labeling system. We have developed a new PSFM, named HTL–Trp–BODIPY–FF, which exhibits strong fluorogenicity upon recognition of Halo-tag protein and reversible fluorescence photoswitching in living cells. This is the first example of a PSFM that can be applicable to a general-purpose Halo-tag protein labeling system for no-wash live-cell imaging.