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TMRE-Mitochondrial Membrane Potential Assay Kit (ab113852) is used for quantifying changes in mitochondrial membrane potential in live cells by flow cytometry, microplate spectrophotometry and fluorescent microscopy.
ab113852 uses TMRE (tetramethylrhodamine, ethyl ester) to label active mitochondria. TMRE is a cell permeant, positively-charged, red-orange dye that readily accumulates in active mitochondria due to their relative negative charge. Depolarized or inactive mitochondria have decreased membrane potential and fail to sequester TMRE.
FCCP (carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone) is a ionophore uncoupler of oxidative phosphorylation. Treating cells with FCCP eliminates mitochondrial membrane potential and TMRE staining. TMRE is suitable for the labeling of mitochondria in live cells and is not compatible with fixation.
|1mM TMRE (in DMSO)||1 x 40µl|
|50mM FCCP (in DMSO)||1 x 10µl|
Our Abpromise guarantee covers the use of ab113852 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Flow Cyt||Use at an assay dependent concentration.|
|FM||Use at an assay dependent concentration.|
P19 neurons (750 cells/mm2) were exposed to MDMA on days 7–9 in serum-free medium for 10 min up to 48 hours. The positive control FCCP (carbonyl cyanide-p-trifluoromethoxyphenylhydrazone), an uncoupler of mitochondrial oxidative phosphorylation, was applied at the concentration of 5 μM for 10 min. The cells were incubated with 500 μM TMRE for 30–45 min at 37°C, 5% CO2, followed by washing once with 100 μl of HBSS containing 0.2% bovine serum albumin. A volume of 200 μL of HBSS containing 0.2% bovine serum albumin was added to each well, and the fluorescence was measured with excitation/emission: 544/590 nm.
Chart showing mean fluorescent intensity +/- standard deviation from quadruplicate measurements of 400 nM TMRE stained Jurkat cells in a 96-well microplate +/- treatment with FCCP.
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