MitoSOX™ Red Mitochondrial Superoxide Indicator, for live-cell imaging

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	- Cells were treated with 5 μM MitoSOX Red for 30 min at room temperature, - Cells were exposed to the β-adrenergic agonist (ISO; 100 nM) and assessed by confocal images.

Protocol tips
- Cells were treated with 5 μM MitoSOX Red for 30 min at room temperature, - Cells were exposed to the β-adrenergic agonist (ISO; 100 nM) and assessed by confocal images.

Publication protocol

Changes in mitochondrial O2*- production were monitored using the fluorescent indicator MitoSOX Red (Invitrogen) and confocal microscopy as described previously [29]. Briefly, cardiomyocytes were loaded with MitoSOX Red (5 μM) for 30 min at room temperature, followed by washout. Cells were exposed to the β-adrenergic agonist (ISO; 100 nM). Confocal images were obtained after 10 min of 1 Hz stimulation in standard Tyrode solution and subsequently after an additional 5 min of 1 Hz stimulation in the presence of ISO. MitoSOX Red fluorescence was measured in the same region of the cell at each time point. The signal from each cell was normalized to that immediately before application of ISO. Changes in ROS induced by application of a high concentration of H2O2 (1 mM) were also monitored using MitoSOX Red. In these experiments the cells were electrically stimulated at 1 Hz for at least 10 min before H2O2 was applied and the fluorescence signal from each cell was normalized to that immediately before application of H2O2.

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