Many redox-active molecules including ROS, reactive nitrogen spec

Many redox-active molecules including ROS, reactive nitrogen species (RNS), and some other redox modifiers (e.g., hydrogen sulfide (H2S)) can diffuse inside cells and across cell membranes, and are increasingly recognized for their important functions in cell signaling [1�C4]. They interact with diverse cellular targets, leading to alterations in their oxidation states and biological functions. In many cases, multiple signaling molecules are generated to interact with the same cellular target in a competitive or synergistic manner. Due to the inherent complexity, a large part of redox homeostasis and signaling remains elusive [4].To examine redox signaling in the cellular and molecular level, a current research focus is to develop methods to identify and characterize molecular products (e.

g., modified macrobiomolecules or small molecule byproducts) resulting from redox biochemical reactions [5]. Another focus is to directly investigate signaling molecules that are actively involved in redox processes. Typically, redox signaling molecules are highly diffusible and reactive, so their detection has been a long-time challenge [6,7]. Colorimetric, electrochemical, and chromatographic assays have been explored. However, these methods often require sample processing, and do not provide much spatial and temporal information about living cells and organisms [8�C10]. In addition, many redox signaling molecules, such as nitric oxide (NO) and peroxynitrite (ONOO?), have very short lifetimes, so it is essentially impossible to directly measure them in processed samples [11,12].

The need to reliably detect redox signaling has promoted the emergence of a group of fluorescent redox probes that can be introduced into living cells and organisms. In previous studies, a large number of synthetic probes have been designed and synthesized [7,13�C16]. These molecules are diverse in structure and show different degrees of sensitivity and selectivity. When loaded into living cells, they could change their fluorescence in response to redox dynamics. Another approach is to use redox probes that Cilengitide are genetically encoded. Genetically encoded probes can be introduced into living cells or organisms in the format of DNA, and next, get expressed into proteins by intracellular machineries. The advantage is that encoded probes can be readily localized to specific cell compartments using corresponding localization sequences or to the vicinity of proteins of interest by creating genetic fusions [17]. Such versatility allows the investigation of biochemical dynamics with subcellular spatial resolution.

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