Real-time monitoring of reactive oxygen and nitrogen species in a multiwell plate using the diagnostic marker products of specific probes.

Developing rigorous assays for cellular detection of reactive oxygen and nitrogen species (O2(·-), H2O2, (·)NO, and ONOO(-)) is an active area of research in our laboratory. Published reports suggest that diagnostic marker products are formed as a result of interaction of these species with small molecular weight fluorescent and nonfluorescent probes. In this chapter, we describe an HPLC-based methodology to detect formation of these species in biological and cellular systems. By monitoring the diagnostic marker products formed from reaction between specific chemical probes and the oxidant species, it is possible to simultaneously assay these species using a multiwell plate (e.g., 384-well plate).