Reactive oxygen and nitrogen species arise in a variety of forms that oxidize, nitrate, nitrosate and halogenate biomolecules. As Balaraman Kalyanaraman at the Medical College of Wisconsin explains, reactive species are critical for the proper functioning of cells. When they go awry, these species can cause an array of diseases ranging from cardiovascular illness to neurodegeneration. But researchers lack real-time and high-throughput methods that clearly identify and quantify the different types of reactive species. In a recent "Paper of the Week" in the Journal of Biological Chemistry, Kalyanaraman and colleagues combined a high-throughput fluorescence plate reader with special probes and high-performance liquid chromatography to detect and quantitate rapidly several reactive species at once. Kalyanaraman says the plate reader works as a first-step screening tool to measure increases in fluorescence caused by superoxide, hydrogen peroxide, nitrosating agents and peroxynitrite. HPLC is then used to confirm and quantify the specific identity of the reactive species. As proof of principle, the investigators demonstrated the real-time monitoring of reactive oxygen and nitrogen species in activated macrophages. They got a better understanding of the rapid sequence of events and their kinetics in the formation of reactive species, processes that have been hard to elucidate in real-time by other approaches. Kalyanaraman says he hopes their method “will lead to more rigorous detection and description of reactive species in all biological models.” He adds that the approach also can be used to screen potential antioxidant and anti-inflammatory drugs rapidly.
Jacek Zielonka, et al.; J. Biol. Chem (2012) doi: jbc.M111.309062
Rajendrani Mukhopadhyay (email@example.com) is the senior science writer for ASBMB Today and the technical editor for the Journal of Biological Chemistry.