Detection of Mitochondrial Mass, Damage, and Reactive Oxygen Species by Flow Cytometry

The reagents and procedures highlighted here will give the investigators an indication of the health status and volume of mitochondria in primary cells and cell lines through the use of a number of cell-permeable dyes. Mitochondrial volume can be monitored by using the probe MitoTracker Green FM. This reagent labels mitochondria in a manner that is independent of the membrane potential, therefore providing a readout relating purely to the mitochondrial mass of the cell. In contrast, MitoTracker Red CMXRos, tetramethylrhodamine methyl ester, and 10-N-nonyl acridine orange label mitochondria in a manner dependent on the membrane potential, thus giving an indication of mitochondrial stress. Using MitoSOX Red, it is also possible to analyze the production of the mitochondrial superoxide anion. Like the MitoTracker probes, MitoSOX Red is taken up passively by cells. In the mitochondria, the probe is oxidized by superoxide, resulting in the emission of red fluorescence.