A Polarity Sensitive Turn-On NIR Fluorescent Probe for Wash-Free Cell Imaging

Wash-free fluorescent probes are highly desirable in bioimaging due to their ability to simplify analytical procedures, shorten detection time, and avoid potential interference or analyte loss caused by washing steps. In this work, we designed and synthesized a donor-π-acceptor (D-π-A) fluorescent molecule, TB-PY, which exhibits strong solvatochromism and a twisted intramolecular charge transfer (TICT) character. Its water-soluble derivative, TBPYS, was subsequently prepared via quaternization of the pyridine unit. TBPYS displays environmentally sensitive near-infrared (NIR) emission, where its fluorescence is effectively quenched in aqueous media but significantly turned on in low-polarity environments due to suppression of the TICT process. This polarity-responsive property was harnessed for wash-free cell imaging. As demonstrated in live Hep-G2 cells, the TBPYS probe produces bright fluorescence in the hydrophobic cytoplasm while remaining dark in the extracellular aqueous environment, enabling high-contrast imaging directly after staining without any washing steps. An outstanding signal-to-noise ratio of 16 was achieved, underscoring the potential of TBPYS as a practical and efficient probe for wash-free bioimaging applications.