Quantifying the Fast Dynamics of HClO in Living Cells by a Fluorescence Probe Capable of Responding to Oxidation and Reduction Events within the Time Scale of Milliseconds.

The biological roles of reactive oxygen species (ROS) depend highly on their dynamics. However, it has been challenging for measuring the dynamics of ROS in cells. In this study, we address a core challenge in developing fluorescence probes for monitoring ROS dynamics by designing a redox couple that can respond rapidly to both oxidation and reduction events. We show that such molecules can be designed by taking advantage of the steric effects of electron-donating groups at the ortho position relative to the selenium center. We demonstrate this design in a new fluorescence probe named Fl-Se. Results reveal that Fl-Se and its oxidized product Fl-SeO rapidly respond to HClO, an important member of the ROS family, and glutathione (GSH), with t1/2 = 2.7 ms at [HClO] = 1 μM; t1/2 = 61 ms at [GSH] = 1 mM. When applied in cells, Fl-Se satisfactorily tracks the dynamics of intracellular HClO in H2O2-stimulated HL-60 cells, as well as the different dynamic behaviors of HClO fluctuations involved in the phorbol 12-myristate-13-acetate-activated immune response of RAW264.7 cells and the 3-deazaneplanocin A-induced apoptosis of HL 60 cells.