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Methods Mol Biol. 2017;1567:15-32. doi: 10.1007/978-1-4939-6824-4_2.

Isolation of Functional Mitochondria from Cultured Cells and Mouse Tissues.

Author information

1
Genzentrum, Department of Biochemistry, Ludwig-Maximilians University, Feodor-Lynen Strasse 25, 81377, Munich, Germany.
2
Institute for Diabetes and Obesity, Helmholtz Zentrum München, 85764, Neuherberg, Germany.
3
Gene Center, LMU Munich and Institute for Diabetes and Obesity, Helmholtz Zentrum München, Munich, Germany. perocchi@genzentrum.lmu.de.

Abstract

Mitochondria serve as the center stage for a number of cellular processes, including energy production, apoptosis, ion homeostasis, iron and copper processing, steroid metabolism, de novo pyrimidine, and heme biosynthesis. The study of mitochondrial function often requires the purification of intact and respiratory-competent organelles. Here, we provide detailed protocols to isolate functional mitochondria from various types of mammalian cells and mouse tissues, in both crude and pure forms. We introduce the use of nitrogen cavitation for the disruption of plasma membrane and the reproducible isolation of mitochondria-enriched fractions of high yield. Mitochondria that are isolated by these procedures are intact and coupled and can directly be used for several downstream analyses, such as measurements of oxygen consumption and calcium buffering capacity.

KEYWORDS:

Crude mitochondria; Mitochondrial integrity; Nitrogen cavitation; Organelle isolation; Pure mitochondria

PMID:
28276010
DOI:
10.1007/978-1-4939-6824-4_2
[Indexed for MEDLINE]

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