Two-photon fluorescence imaging of mitochondrial viscosity with water-soluble pyridinium inner salts

Rational design of mitochondrial viscosity probes with water-soluble two-photon (TP) fluorescence probes still remains a challenge in practical biological applications. Herein, three novel water-soluble TP active sulfosalt derivatives (Mito-Z, Mito-H, and Mito-Q) were developed with high sensitivity to mitochondrial viscosity in living cells. In non-viscous media, the probes possess a small fluorescence brightness (ϵΦ) and TP action cross-sections (Φδ). With the increasing viscosity of the micro-environment, ϵΦ and Φδ of the probes increase significantly. Moreover, the results of TP fluorescence imaging demonstrated that Mito-Q was sensitive to local viscosity and selectively staining mitochondria. In a nutshell, Mito-Q possessed a number of beneficial properties in bioimaging such as good membrane permeability, long wavelength red emission, moderate Φδ value, high sensitivity viscosity response, and specific mitochondrial targeting, affording a promising platform for detecting mitochondrial viscosity.