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Nat Protoc. 2008;3(6):965-76. doi: 10.1038/nprot.2008.61.

Analysis of mitochondrial function in situ in permeabilized muscle fibers, tissues and cells.

Author information

  • 1Daniel Swarovski Research Laboratory, Department of General and Transplant Surgery, Innsbruck Medical University, Innsbruck A6020, Austria. andrey.kuznetsov@uki.at

Abstract

Analysis of mitochondrial function is central to the study of intracellular energy metabolism, mechanisms of cell death and pathophysiology of a variety of human diseases, including myopathies, neurodegenerative diseases and cancer. However, important properties of mitochondria differ in vivo and in vitro. Here, we describe a protocol for the analysis of functional mitochondria in situ, without the isolation of organelles, in selectively permeabilized cells or muscle fibers using digitonin or saponin. A specially designed substrate/inhibitor titration approach allows the step-by-step analysis of several mitochondrial complexes. This protocol allows the detailed characterization of functional mitochondria in their normal intracellular position and assembly, preserving essential interactions with other organelles. As only a small amount of tissue is required for analysis, the protocol can be used in diagnostic settings in clinical studies. The permeabilization procedure and specific titration analysis can be completed in 2 h.

PMID:
18536644
[PubMed - indexed for MEDLINE]
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