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12664
Mitochondrial Membrane Potential Assay Kit (I)

Mitochondrial Membrane Potential Assay Kit (I) #12664

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Flow Cytometry Image 1

Figure 1. Flow cytometric analysis of Jurkat cells labeled with JC-1 (2 μM), untreated (red) or CCCP-treated (50 μM, 37ºC, 30 min; blue).

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Fluorescent Detection Image 2

Figure 2. Confocal immunofluorescent analysis of NIH/3T3 cells seeded at 2x105 cell/ml in 96-well black plate with clear bottom and incubated overnight. Cells were then treated with JC-1 (2 μM, 30 min; left) or with JC-1 (2 μM, 30 min) and CCCP (400 μM, 30 min; right).

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Fluorescent Detection Image 3

Figure 3. HeLa (3x105 cell/ml, green) and NIH/3T3 (2x105 cell/ml, orange) cells were incubated with JC-1 (2 μM, 30 min) and various concentrations of CCCP, as indicated. Relative fluorescent units (RFU) are shown on the left, while the RFU Ratio (RFU(red)/RFU(green)) is shown on the right.

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Product Includes Quantity (with Count) Solution Color
JC-1 5 x 15 µg Red
CCCP 1 x 100 µl Yellow
Phosphate Buffered Saline (PBS-20X) 9808 1 x 25 ml Colorless

Mitochondrial Membrane Potential Assay Kit (I) is a fluorescent assay that detects the mitochondrial membrane potential in living cells. The kit includes the cationic dye JC-1 (5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethyl-imidacarbocyanine iodide) and a mitochondrial membrane potential disruptor CCCP (carbonyl cyanide 3-chlorophenylhydrazone). JC-1 is a cell membrane permeable, fluorescent dye with green emission (~520 nm). When JC-1 accumulates in intact mitochondria, the dye forms aggregates that lead to orange-red fluorescence (~590 nm). The relative fluorescent units (RFU) of the orange-red emission, and the ratio of red to green emission, can be used as an indicator for mitochondria membrane potential.

Mitochondrial Membrane Potential Assay Kit (I) is expected to detect the mitochondrial membrane potential in living cells across all species. For the best result, a cell number titration is recommended when using a plate-reader and a 96-well plate.

Species Reactivity:

All Species Expected

Mitochondria function as the main cellular powerhouse and play important roles in other processes, such as steroid metabolism, calcium homeostasis, apoptosis, and cellular proliferation. Mitochondrial membrane potential is a key indicator of mitochondrial function and cell health (1,2). The dissipation of mitochondrial membrane potential is considered an early indicator of apoptosis (3).

JC-1 (5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethyl-imidacarbocyanine iodide) is a cell membrane permeable, cationic dye. In normal cells, JC-1 concentrates in mitochondria to form aggregates in response to relatively high membrane potential. Decreased mitochondrial membrane potential results in dispersal of mostly monomeric JC-1 throughout the cell. When excited at 490 nm, JC-1 monomers emit a green fluorescence with a maximum at ~520 nm. Aggregates of JC-1 emit an orange-red fluorescence with a maximum at ~590 nm. Therefore, the fluorescence intensity of the orange-red emission and the ratio of orange-red fluorescence to green fluorescence can be used to measure mitochondrial membrane potential and serve as an indicator of overall cell health (4).

  1. Perry, S.W. et al. (2011) Biotechniques 50, 98-115.
  2. Nesti, C. et al. (2007) Biosci Rep 27, 165-71.
  3. Petit, P.X. et al. (1995) J Cell Biol 130, 157-67.
  4. Perelman, A. et al. (2012) Cell Death Dis 3, e430.
For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

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