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

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.