What is MitoSOX Red?

MitoSOX™ Red reagent is a novel fluorogenic dye specifically targeted to mitochondria in live cells. Oxidation of MitoSOX™ Red reagent by superoxide produces red fluorescence. • Readily oxidized by superoxide but not by other ROS- or RNS-generating systems. • Absorption/emission maxima: ∼510/580 nm.

What does MitoSOX detect?

MitoSOX-based assays are widely used to detect mitochondrial reactive oxygen species (ROS), especially superoxide. To this end, 5 μM MitoSOX is commonly used.

Can MitoSOX be fixed?

the uptake of MitoSOX into the mitochondria is dependent on the mitochondrial membrane potential. When there is no membrane potential anymore, MitoSOX will leak out. Hence, you can’t use MitoSOX on fixed cells.

How do mitochondria produce ROS?

Generation of mitochondrial ROS mainly takes place at the electron transport chain located on the inner mitochondrial membrane during the process of oxidative phosphorylation. Collectively, both superoxide and hydrogen peroxide generated in this process are considered as mitochondrial ROS.

How do you find ROS?

Many means are available for ROS detection, such as the classical and commonly used spectrophotometry methods, use of fluorescence and chemiluminescence probes, and electron spin resonance (ESR/EPR), which provide direct identification of different types of oxygen radicals.

How do you measure ROS levels?

Generally, it is a good idea to measure ROS by two independent methods and monitor different species in the same samples. A wide variety of spin traps and spin probes such as DMPO, PBN, TEMPOL, and mito-TEMPO protect the brain, heart, and vascular tissue under conditions of oxidative stress (8, 19, 64).

Are there any positive controls for mitosox red superoxide detection?

Knockdown of the mitochondrial fluorescence signal by SOD mimetics such as FeTCPP and the manganese porphyrin MnTBAP provides valuable negative controls in MitoSOX™ Red superoxide detection assays. Useful positive controls (not shown) include treatment with antimycin A, doxorubicin, or high glucose.

How are mitosox red reagents used in live cells?

MitoSOX™ Red reagent is a novel fluorogenic dye specifically targeted to mitochondria in live cells. Oxidation of MitoSOX™ Red reagent by superoxide produces red fluorescence. The production of superoxide by mitochondria can be visualized in fluorescence microscopy using the MitoSOX™ Red reagent.

How is mitosox red indicator used in flow cytometry?

Detection of MitoSOX™ Red indicator by flow cytometry typically uses the FL2 emission channel (585/42 nm). A common application of MitoSOX™ Red indicator is the correlation of superoxide generation with the accumulation of oxidative damage products such as lipid peroxides and modified DNA bases such as 8-oxodG.

How is mitosox used to diagnose mitochondrial dysfunction?

Cells were then labeled with MitoSOX™ Red indicator in combination with blue-fluorescent Hoechst 33342 nuclear stain. Knockdown of the mitochondrial fluorescence signal by SOD mimetics such as FeTCPP and the manganese porphyrin MnTBAP provides valuable negative controls in MitoSOX™ Red superoxide detection assays.