Environmentally sensitive fluorescent probes for the visualization of selective proteins: Recent progress and...

Environmentally sensitive fluorescent probes represent an emerging class of chemical tools that enable the visualization of selective proteins. These probes typically consist of a target-selective ligand, an appropriate linker and a responsive fluorophore that generally undergoes turn-on emission when its local microenvironments such as polarity and viscosity changes. In contrast to traditional always-on dyes, this modular design affords high signal-to-noise ratios and wash-free imaging. Recent advances have expanded the application of such probes from kinases and histone deacetylases to regulators of protein-protein interaction, metabolic enzymes, channels and receptors. Key strategies for the design of these probes include the use of environmentally sensitive fluorophores to enhance fluorescence contrast. A couple of turn-on probes have been applied to assess the activity of proteolysis-targeting chimeras (PROTACs), while fluorescent PROTACs allow direct tracking of target degradation in living systems. This review summarizes recent advances (2020-2025) in strategies for design, mechanisms of activation, and biological applications, and outlines future directions including the development of NIR-II fluorophores, AI-assisted discovery and in vivo translation.