Real-time measurement of intracellular reactive oxygen species using mito tracker orange (CMH2TMRos)

被引:26
作者
Kweon, SM
Kim, HJ
Lee, ZW
Kim, SJ
Kim, SI
Paik, SG
Ha, KS
机构
[1] Korea Basic Sci Inst, Biomol Res Team, Yusung Ku, Taejon 305333, South Korea
[2] Chungnam Natl Univ, Dept Biol, Taejon 305764, South Korea
关键词
reactive oxygen species; arachidonic acid; lysophosphatidic acid; MitoTracker orange; real-time measurement;
D O I
10.1023/A:1013290316939
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
We have investigated a novel method to monitor real changes of intracellular ROS by the use of CMH(2)TMRos (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 CMH(2)TMRos. 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 CMH(2)TMRos is a useful fluorophore for real-time monitoring of intracellular ROS and also indicated that CMH(2)TMRos detects primarily superoxide in cells even though the fluorophore can be oxidized by both superoxide and H2O2.
引用
收藏
页码:341 / 352
页数:12
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