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Published on: October 3, 2012
Mitochondrial Morphofunctional Profiling in Primary Human Skin Fibroblasts Using TMRM and Mitotracker Green
Jesper M M Bergmans1, Els M A van de Westerlo2, Sander Grefte3
1Department of Pediatrics, Amalia Children's Hospital, Radboud Center for Mitochondrial Medicine, Radboud University Medical Center, Nijmegen, The Netherlands.
Abstract:
Mitochondrial morphology and membrane potential (Δψ) are important readouts of mitochondrial function. Integrated analysis of these parameters in living cells can be performed using fluorescent lipophilic cations, which enter cells and accumulate in the mitochondrial matrix in a Δψ-dependent manner. Here, we describe the use of tetramethylrhodamine methyl ester (TMRM) and Mitotracker Green FM (MG) for mitochondrial morphology and semiquantitative Δψ analysis in living primary human skin fibroblasts (PHSFs). Practically, we present an integrated protocol to quantify mitochondrial morphology parameters and signal intensity using epifluorescence microscopy of PHSFs co-stained with TMRM and MG. This approach performs best using large flat cells like PHSFs, which display a high mitochondria-specific fluorescence signal and are imaged at a relatively high (x40) magnification.
