Recent Advances in Molecular Engineering of Organelle-Targeted Viscosity Probes for Fluorescence and Biological...

Eukaryotic cell function relies on tightly regulated organelle microenvironments, where local viscosity-a key biophysical parameter reflecting molecular crowding-directly governs processes like molecular diffusion, enzymatic reactions, and metabolite transport. The dynamic balance of viscosity is essential for cellular homeostasis, while dysregulation of organellar viscosity is a critical hallmark of diseases such as lysosomal storage disorders, neurodegeneration, and cancer. However, conventional imaging lacks the spatiotemporal resolution for dynamic, subcellular viscosity monitoring. This gap has been addressed by advanced fluorescent probes, which integrate organelle-targeting units (e.g., triphenylphosphonium for mitochondria, morpholine for lysosomes) with viscosity-sensitive fluorophores, often based on molecular rotors. These tools accumulate selectively within organelles, converting viscosity changes into quantifiable fluorescence signals and enabling real-time, high-resolution imaging. This review systematically examines the design principles, targeting mechanisms, and biological applications of viscosity-responsive probes for lysosomes, mitochondria, the endoplasmic reticulum, and the Golgi apparatus while exploring mechanistic links between aberrant viscosity and disease. These probes serve as powerful tools for fundamental cell biology and for establishing a technical foundation in early diagnosis, mechanistic elucidation, and targeted therapy. The next generation of probes, integrating multiparameter detection, improved tissue penetration, and enhanced biocompatibility, holds strong promise for transforming the field and delivering deeper insights into cellular function and disease.