Universität zu Köln

Franko, Andras ORCID: 0000-0003-0485-5439, Baris, Olivier R., Bergschneider, Eva, von Toerne, Christine ORCID: 0000-0002-4132-4322, Hauck, Stefanie M., Aichler, Michaela, Walch, Axel K., Wurst, Wolfgang ORCID: 0000-0003-4422-7410, Wiesner, Rudolf J., Johnston, Ian C. D. and de Angelis, Martin Hrabe ORCID: 0000-0002-7898-2353 (2013). Efficient Isolation of Pure and Functional Mitochondria from Mouse Tissues Using Automated Tissue Disruption and Enrichment with Anti-TOM22 Magnetic Beads. PLoS One, 8 (12). SAN FRANCISCO: PUBLIC LIBRARY SCIENCE. ISSN 1932-6203

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Abstract

To better understand molecular mechanisms regulating changes in metabolism, as observed e. g. in diabetes or neuronal disorders, the function of mitochondria needs to be precisely determined. The usual isolation methods such as differential centrifugation result in isolates of highly variable quality and quantity. To fulfill the need of a reproducible isolation method from solid tissues, which is suitable to handle parallel samples simultaneously, we developed a protocol based on anti-TOM22 (translocase of outer mitochondrial membrane 22 homolog) antibody-coupled magnetic beads. To measure oxygen consumption rate in isolated mitochondria from various mouse tissues, a traditional Clark electrode and the high-throughput XF Extracellular Flux Analyzer were used. Furthermore, Western blots, transmission electron microscopic and proteomic studies were performed to analyze the purity and integrity of the mitochondrial preparations. Mitochondrial fractions isolated from liver, brain and skeletal muscle by anti-TOM22 magnetic beads showed oxygen consumption capacities comparable to previously reported values and little contamination with other organelles. The purity and quality of isolated mitochondria using anti-TOM22 magnetic beads was compared to traditional differential centrifugation protocol in liver and the results indicated an obvious advantage of the magnetic beads method compared to the traditional differential centrifugation technique.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Franko, Andras UNSPECIFIED orcid.org/0000-0003-0485-5439 UNSPECIFIED Baris, Olivier R. UNSPECIFIED UNSPECIFIED UNSPECIFIED Bergschneider, Eva UNSPECIFIED UNSPECIFIED UNSPECIFIED von Toerne, Christine UNSPECIFIED orcid.org/0000-0002-4132-4322 UNSPECIFIED Hauck, Stefanie M. UNSPECIFIED UNSPECIFIED UNSPECIFIED Aichler, Michaela UNSPECIFIED UNSPECIFIED UNSPECIFIED Walch, Axel K. UNSPECIFIED UNSPECIFIED UNSPECIFIED Wurst, Wolfgang UNSPECIFIED orcid.org/0000-0003-4422-7410 UNSPECIFIED Wiesner, Rudolf J. UNSPECIFIED UNSPECIFIED UNSPECIFIED Johnston, Ian C. D. UNSPECIFIED UNSPECIFIED UNSPECIFIED de Angelis, Martin Hrabe UNSPECIFIED orcid.org/0000-0002-7898-2353 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-470156
DOI:	10.1371/journal.pone.0082392
Journal or Publication Title:	PLoS One
Volume:	8
Number:	12
Date:	2013
Publisher:	PUBLIC LIBRARY SCIENCE
Place of Publication:	SAN FRANCISCO
ISSN:	1932-6203
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language DENSITY GRADIENT CENTRIFUGATION; SKELETAL-MUSCLE; LABEL-FREE; RAT-BRAIN; DYSFUNCTION; MICE; HOMEOSTASIS; DISORDERS; ORGANELLE; NEURONS Multiple languages Multidisciplinary Sciences Multiple languages
URI:	http://kups.ub.uni-koeln.de/id/eprint/47015