Construction of fluorescent rotors with multiple intramolecular rotation sites for visualization of cellular viscous compartments with elevated fidelity.

Viscosity-sensitive fluorescent dyes are widely utilized to image viscous intracellular compartments with high fidelity. However, the sensitivity of many fluorescent rotors needs improvement for bioimaging applications. Herein, we proposed to construct a fluorescent rotor with multiple intramolecular rotation sites to elevate its sensitivity to environmental viscosity. The fabricated fluorescent rotor showed evidently increased sensitivity to viscosity and had the potential to image intracellular viscous compartments with improved fidelity. By decorating the rotor with sidechains of different lengths, we successfully fabricated two fluorescent probes, TAPI-6 and TAPI-16, to visualize the mitochondria and plasma membrane, respectively, with high fidelity. The two probes were also successfully utilized to clearly visualize the mitochondria and plasma membranes in skeletal muscle tissue, cardiac muscle tissue, and liver tissue, demonstrating the potential of the fluorescent rotor for bioimaging applications. We believe that the strategy of increasing the sensitivity to viscosity using multiple rotation sites is valuable for the construction of fluorescent rotors, and the presented fluorescent probes in this work can serve as powerful tools for biological research.