We have investigated a novel method to monitor real changes of intracellular ROS by the use of CMH2TMRos (a reduced form of MitoTracker orange) in Swiss 3T3 fibroblasts. Arachidonic acid induced a rapid increase of CMTMRos fluorescence with a maximal elevation at 120–150 sec, which was determined by scanning every 10 sec with a confocal microscope. The fluorescence increase by arachidonic acid was completely inhibited by 2-MPG but not by catalase, indicating a major contribution of superoxide to the oxidation of CMH2TMRos. Incubation with glucose oxidase, exogenous H2O2, KO2 and lysophosphatidic acid also increased the CMTMRos fluorescence, which was blocked by 2-MPG. These results suggested that CMH2TMRos is a useful fluorophore for real-time monitoring of intracellular ROS and also indicated that CMH2TMRos detects primarily superoxide in cells even though the fluorophore can be oxidized by both superoxide and H2O2.
Real-Time Measurement of Intracellular Reactive Oxygen Species Using Mito Tracker Orange (CMH2TMRos)
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Soo-Mi Kweon, Hyun-Jung Kim, Zee-Won Lee, Soo-Jung Kim, Seung-IL Kim, Sang-Gi Paik, Kwon-Soo Ha; Real-Time Measurement of Intracellular Reactive Oxygen Species Using Mito Tracker Orange (CMH2TMRos). Biosci Rep 1 June 2001; 21 (3): 341–352. doi: https://doi.org/10.1023/A:1013290316939
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