Construction of a benzindole-based fluorescent probe with high anti-interference capability for viscosity sensing in complex environments

Viscosity, a key parameter regulating the cellular microenvironment, is closely linked to various complex physiological and pathological processes. In the design of viscosity probes, indole groups are often selected as electron-withdrawing groups due to their suitable electronic properties. However, traditional indole-based viscosity-responsive fluorescent probes have poor stability, small Stokes shifts, and weak anti-interference, hampering specific detection of viscosity changes in complex cellular microenvironments. To address these limitations, we optimized the structure of the traditional indole group and report a p-benzindole-based viscosity fluorescence probe MTPA-WY, which exhibits excellent viscosity-responsive optical properties, with a fluorescence intensity enhancement of approximately 30-fold at 657 nm, a large Stokes shift up to 131 nm, and excellent anti-interference capability against HClO, H₂S. Additionally, cytotoxicity assays and colocalization experiments confirmed the probe's excellent biocompatibility and mitochondrial targeting. This probe enables selective monitoring of viscosity fluctuations and long-term dynamic imaging in living cells, while demonstrating exceptional resistance to interference from various microenvironmental interferents, such as HClO, H₂S and other reactive species. Compared to traditional indole-based probes, MTPA-WY overcomes inherent limitations through structural optimization and provides a new tool for studying viscosity-related disease mechanisms in the mitochondrial microenvironment.