pH-Switchable Fluorescent Probe for Spatially-Confined Visualization of Intracellular Hydrogen Peroxide

Intracellular H2O2 plays an important role in regulating a variety of cellular functions. Fluorescent probes that can make response to intracellular levels of H2O2 would provide valuable tools for revealing the functions of H2O2 in living organisms. However, traditional pH-insensitive probes and lysosome-targetable probes can only provide spatially nonspecific visualization of intracellular H2O2 and specific sensing of lysosomal H2O2, respectively. In this work, we developed a H2O2-responsive and pH-switchable fluorescent probe (HP-L1) which can make response sequentially to intracellular H2O2 and lysosomal pH. The fluorescent probe is comprised of a H2O2-responsive boronate moiety and a pH-switchable spirobenzopyran fluorophore. When the probe was applied for intracellular H2O2 sensing, only fluorescent emission from lysosomes is visible, and the fluorescence from other regions is not able to be obviously detected, which is due to the pH-switchable property of the spirobenzopyran fluorophore. Thus, the developed fluorescence probe enables the spatially confined (i.e., lysosome-specific) visualization of the intracellular H2O2. We envisioned that this kind of fluorescent probe (or the proposed sensing strategy) would allow the visualization of the overall levels of intracellular H2O2 without interferences of possible fluorescent signals from other sources (e.g., dyes for cellular staining and multiplex analysis).