FLIM-guided self-reporting of PDT efficacy using a mitochondria-targeted, viscosity-sensitive quinolinium probe

Mitochondrial viscosity plays a pivotal role in the microenvironment of mitochondria and is closely associated with pathological conditions. As a light-activated treatment for cancer and infections, photodynamic therapy (PDT) relies on reactive oxygen species (ROS) generation by photosensitizers to induce cell apoptosis, and mitochondria are prime targets of PDT. However, current photosensitizers lack real-time feedback on therapeutic efficacy. Herein, we report Mito-Qu and Mito-QuE, two quinolinium-based multifunctional fluorescent probes for mitochondrial targeting and viscosity response based on the D-π-A structure to overcome these limitations. Mito-Qu and Mito-QuE reveal exceptionally large Stokes shifts of 179 nm and 186 nm and exhibit viscosity-sensitive emission via a twisted intramolecular charge transfer (TICT) mechanism, making them robust sensors for monitoring mitochondrial viscosity dynamics. Additional experiments suggest that Mito-Qu acts as a sensitive monitor for mitochondrial viscosity monitoring and self-reporting PDT efficacy via both confocal imaging and FLIM, providing a blueprint for developing next-generation organelle-targeted probes for intracellular microenvironment monitoring.