Send to

Choose Destination

See 1 citation found by title matching your search:

PLoS One. 2018 Apr 26;13(4):e0196632. doi: 10.1371/journal.pone.0196632. eCollection 2018.

Isolation of mitochondria from Saccharomyces cerevisiae using magnetic bead affinity purification.

Author information

Department of Pathology and Cell Biology, Columbia University, New York, NY, United States of America.
Department of Neurology, Columbia University, New York, NY, United States of America.
Department of Psychiatry, University of Pittsburgh, Pittsburgh, PA, United States of America.
Department of Cell Biology, University of Pittsburgh, Pittsburgh, PA, United States of America.


Isolated mitochondria are widely used to study the function of the organelle. Typically, mitochondria are prepared using differential centrifugation alone or in conjunction with density gradient ultracentrifugation. However, mitochondria isolated using differential centrifugation contain membrane or organelle contaminants, and further purification of crude mitochondria by density gradient ultracentrifugation requires large amounts of starting material, and is time-consuming. Mitochondria have also been isolated by irreversible binding to antibody-coated magnetic beads. We developed a method to prepare mitochondria from budding yeast that overcomes many of the limitations of other methods. Mitochondria are tagged by insertion of 6 histidines (6xHis) into the TOM70 (Translocase of outer membrane 70) gene at its chromosomal locus, isolated using Ni-NTA (nickel (II) nitrilotriacetic acid) paramagnetic beads and released from the magnetic beads by washing with imidazole. Mitochondria prepared using this method contain fewer contaminants, and are similar in ultrastructure as well as protein import and cytochrome c oxidase complex activity compared to mitochondria isolated by differential centrifugation. Moreover, this isolation method is amenable to small samples, faster than purification by differential and density gradient centrifugation, and more cost-effective than purification using antibody-coated magnetic beads. Importantly, this method can be applied to any cell type where the genetic modification can be introduced by CRISPR or other methods.

[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Public Library of Science Icon for PubMed Central
Loading ...
Support Center