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Methods Mol Biol. 2019;1928:69-76. doi: 10.1007/978-1-4939-9027-6_5.

Measurement of Mitochondrial Membrane Potential with the Fluorescent Dye Tetramethylrhodamine Methyl Ester (TMRM).

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Monash Micro Imaging, Hudson Institute of Medical Research, Clayton, VIC, Australia.
Centre for Innate Immunity and Infectious Diseases, Hudson Institute of Medical Research, Clayton, VIC, Australia.
Department of Molecular and Translational Science, Monash University, Clayton, VIC, Australia.
School of Life and Environmental Sciences, Deakin University, Waurn Ponds, VIC, Australia.


The mitochondrial membrane potential (Δψm) drives the generation of ATP by mitochondria. Interestingly, Δψm is higher in many cancer cells comparted to healthy noncancerous cell types, providing a unique metabolic marker. This feature has also been exploited for therapeutic use by utilizing drugs that specifically accumulate in the mitochondria of cancer cells with high Δψm. As such, the assessment of Δψm can provide very useful information as to the metabolic state of a cancer cell, as well as its potential for malignancy. In addition, the measurement of Δψm can also be used to test the ability of novel anticancer therapies to disrupt mitochondrial metabolism and cause cell death.Here, we outline two methods for assessing Δψm in cancer cells using confocal microscopy and the potentiometric fluorescent dye tetramethylrhodamine methyl ester (TMRM). In the first protocol, we describe a technique to quantitatively measure Δψm, which can be used to compare Δψm between different cell types. In the second protocol, we describe a technique for assessing changes to Δψm over time, which can be used to determine the effectiveness of different therapeutic compounds or drugs in modulating mitochondrial function.


Cancer cells; Confocal imaging; Fluorescence; Membrane potential; Mitochondria; Osteosarcoma; TMRM

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