Simple quantitative detection of mitochondrial superoxide production in live cells

Biochem Biophys Res Commun. 2007 Jun 22;358(1):203-8. doi: 10.1016/j.bbrc.2007.04.106. Epub 2007 Apr 25.


Experiments with isolated mitochondria have established that these organelles are pivotal intracellular sources of superoxide in a variety of pathophysiological conditions. Recently, a novel fluoroprobe MitoSOX Red was introduced for selective detection of superoxide in the mitochondria of live cells and was validated with confocal microscopy. Here we show approximately 3-7 fold dose- and time-dependent increase in mitochondrial superoxide production (measured by MitoSOX using flow cytometry and confocal microscopy) in rat cardiac derived H9c2 myocytes and/or in human coronary artery endothelial cells triggered by Antimycin A, Paraquat, Doxorubicin or high glucose. These results establish a novel, quantitative method for simple detection of mitochondrial superoxide generation simultaneously in a large population of live cells by flow cytometry. This method can also be adapted for immune cell studies with mixed population of T or B cells or their subsets to analyze mitochondrial superoxide levels using multiple labeled surface markers in individual populations.

Publication types

  • Research Support, N.I.H., Intramural

MeSH terms

  • Animals
  • Antimycin A / pharmacology
  • Cells, Cultured
  • Coronary Vessels / cytology
  • Doxorubicin / pharmacology
  • Endothelial Cells / chemistry*
  • Endothelial Cells / drug effects
  • Endothelium, Vascular / cytology
  • Endothelium, Vascular / drug effects
  • Flow Cytometry
  • Fluorescent Dyes
  • Glucose / pharmacology
  • Humans
  • Microscopy, Confocal
  • Mitochondria, Heart / chemistry*
  • Mitochondria, Heart / drug effects
  • Myocytes, Cardiac / chemistry*
  • Myocytes, Cardiac / drug effects
  • Paraquat / pharmacology
  • Phenanthridines
  • Rats
  • Superoxides / analysis*


  • 5-(6'-triphenylphosphoniumhexyl)-5,6-dihydro-6-phenyl-3,8-phenanthridinediammine
  • Fluorescent Dyes
  • Phenanthridines
  • Superoxides
  • Antimycin A
  • Doxorubicin
  • Glucose
  • Paraquat