After cells were seeds onto a 24-well plate at a density of 105 cells/well, mitochondrial membrane potential was measured as described previously [34 (link)35 (link)]. Briefly, at the indicated time points, cells were incubated with 200 nM tetramethylrhodamine ethyl ester perchlorate (TMRE) in 0.1% DMSO for 20 minutes at 37℃. After washing six times with PBS, 1% Triton X-100 was added to each well. TMRE intensity for 200 µl cell lysate on a Nunc 96-well black plate (Thermo Fisher Scientific) was then quantified with the SpectraMax i3 plate reader (Molecular Devices) using 549 nm for excitation and 575 nm for emission.
Protocol detail
Quantifying Mitochondrial Membrane Potential
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Cell
Devices
Dmso
Mitochondrial membrane potential
Perchlorate
Seeds
Tetramethylrhodamine ethyl ester
Triton x 100
Corresponding Organization : Jeju National University
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Variable analysis
independent variables
- Cell seeding density (10^5 cells/well)
- Mitochondrial membrane potential
- Time points
- TMRE concentration (200 nM)
- DMSO concentration (0.1%)
- Incubation time (20 minutes)
- Temperature (37°C)
- Number of PBS washes (6 times)
- Triton X-100 concentration (1%)
- Plate type (Nunc 96-well black plate)
- Plate reader (SpectraMax i3)
- Excitation wavelength (549 nm)
- Emission wavelength (575 nm)
- Positive control: Not explicitly mentioned
- Negative control: Not explicitly mentioned
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