Abstract
Detecting biothiols like glutathione (GSH), homocysteine (Hcy), and cysteine (Cys) is key to understanding their roles in health and disease. We developed BT-DNBS, a cyanine-based turn-on fluorescent probe with a dinitrobenzenesulfonyl (DNBS) quencher group. Upon biothiol interaction, the quencher is cleaved, restoring fluorescence. The resulting probe BT-NH shows a maximum emission wavelength at 630 nm and a large Stokes shift (≈200 nm), enhancing detection accuracy. Low cytotoxicity and high time resolution make BT-DNBS suitable for live-cell imaging. Imaging of A431 cells confirmed intracellular biothiol detection, with NEM pre-treatment reducing fluorescence, verifying specificity. BT-DNBS holds promise for biomedical research, particularly in disease diagnostics.