Figure 1. Flow cytometric analysis of Jurkat cells, unlabeled (black) or labeled with 200 nM TMRE and treated with 0 μM CCCP (blue), 3.2 μM CCCP (green), or 80 μM CCCP (red).
Figure 2. Confocal immunofluorescent analysis of NIH/3T3 cells (2x105 cell/ml) seeded in a 96-well black plate with a clear bottom and incubated overnight. Cells were untreated or treated with CCCP (400 μM, 20 min) followed by labeling with TMRE (200 nM, 30 min).
Figure 3. HeLa cells (3x105 cell/ml) were treated with various concentrations of CCCP for 15 minutes prior to labeling with 200 nM TMRE.
The Mitochondrial Membrane Potential Assay Kit (II) is a fluorescent assay that detects the mitochondrial membrane potential in living cells. The kit includes the cationic dye TMRE (tetramethylrhodamine ethyl ester perchlorate) and a mitochondrial membrane potential disruptor CCCP (carbonyl cyanide 3-chlorophenylhydrazone). TMRE is a cell membrane permeable, fluorescent dye that accumulates in intact mitochondria. Depolarized or inactive mitochondria exhibit decreased membrane potential, resulting in reduced TMRE accumulation.
The Mitochondrial Membrane Potential Assay Kit (II) is expected to detect the mitochondrial membrane potential in living cells cross all species. For the best result, a cell number titration is recommended when using a plate-reader and a 96-well plate.
Mitochondria are the main power house in cells and play important roles in processes such as steroid metabolism, calcium homeostasis, apoptosis and cellular proliferation. Mitochondrial membrane potential is a key indicator of its function and cell health (1,2). The dissipation of mitochondrial membrane potential is established as an early indicator for apoptosis (3).
TMRE (tetramethylrhodamine, ethyl ester) is a cell membrane permeable cationic dye. In normal cells, TMRE accumulates in the mitochondria in response to their high membrane potential and negative charge. When excited at 550 nm, TMRE emits an orange-red fluorescence with a maximum at 575 nm (orange-red). Cells that have lost membrane potential or mitochondria activity cannot accumulate TMRE. Therefore, the fluorescence intensity of the orange-red emission can be used to measure mitochondria membrane potential and is an indicator for cell health (4).
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