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We have developed a new reversible fluorescence probe MPhSe-BOD for the redox cycle process between hypochlorous acid and hydrogen sulfide in solution and in living cells. Confocal microscopy imaging using RAW264.7 cell lines shows that the probe has good cell membrane permeability, and can monitor intracellular HClO/H 2 S redox cycles continuously.

A diselenide containing fluorescent probe based on a fluorescein scaffold for thiols was developed. The fluorescent probe exhibited rapid response, high selectivity and reversibility. Confocal fluorescence microscopy was used to visualize the redox changes mediated by thiols and reactive oxygen species in living HeLa cells.Intracellular thiols play an essential role in maintaining redox homeostasis by regulating the redox status between reduced free thiols and oxidized disulfides.1 Glutathione (GSH), which has been identified as the most abundant non-protein thiol, is considered to be the main player in combating oxidative stress and regulating the redox environment of the internal cellular compartments. Further development of intracellular thiol detection methods is necessary to understand the complicated roles of thiols in redox biology and elucidate the pathogenesis of various disorders in living systems. Compared with other approaches, fluorescence imaging is less invasive, more sensitive and more convenient. 11Numerous fluorescent probes for thiols based on Michael addition, aldehyde cyclization, fluorophore deprotection, and thiol-disulfide exchange protocols have been reported. 2,12 However, very few of these probes react with thiols rapidly enough to achieve real-time detection in vivo. And most of these probes can only respond to thiols statically. Research has shown that fluorescent probes which are capable of responding to thiols and ROS reversibly offer better chances of visualizing redox signaling, stress, and thiols repair. Real-time methods for measuring thiols in vivo based on redoxsensitive green fluorescent protein have been reported. 1,14 However, such techniques require gene manipulation while redox-responsive probes based on small molecules are scarce. 13,15Glutathione peroxidase (GPx) is a selenoenzyme that protects various organisms from oxidative damage by catalyzing ROS reduction in the presence of GSH. 16 To clarify the catalytic mechanism of GPx and develop antioxidant drugs, researchers have synthesized many organoselenium compounds as GPx mimics, in which monoselenides, diselenides, and ebselen analogs are the major ones. 16,17 Steinmann et al. 18 recently reported that the cleavage of diselenide bonds by thiols is 5 orders of magnitude faster than that of disulfide bonds. Building on these results, we designed and synthesized a diselenide-based fluorescent probe (FSeSeF, Scheme 1) for the rapid detection of thiols and redox changes mediated by thiols and ROS in living cells. FSeSeF is a weakly fluorescent molecule that contains a diselenide bond as a fast, reversible recognition center for thiols and two fluoresceins as signal reporters. Once FSeSeF is cleaved by GSH, both the selenenyl sulfide FSeSG and the Scheme 1 Structure of FSeSeF and the nucleophilic attack by GSH at Se-Se bond with the generation of selenenyl sulfide FSeSG and selenol FSeH.