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Isolation and Functional Analysis of Mitochondria from Cultured Cells and Mouse Tissue
Published on: March 23, 2015
Isolation of mitochondria from the CNS
1Department of Anesthesiology, Organized Research Center, School of Medicine, University of Maryland, Baltimore, Maryland, USA.
Current Protocols in Neuroscience
|June 26, 2010
Summary
This study details a protocol for isolating brain mitochondria using Percoll gradient centrifugation. The method effectively separates various cellular components, yielding purified mitochondria with excellent respiratory function.
Area of Science:
- Neuroscience
- Cell Biology
- Biochemistry
Background:
- Mitochondria are crucial for cellular energy production.
- Accurate isolation of brain mitochondria is essential for studying neurological functions and diseases.
- Existing methods may yield contaminated or damaged mitochondrial fractions.
Purpose of the Study:
- To describe a protocol for the isolation of pure and functional brain mitochondria.
- To separate synaptic and nonsynaptic mitochondria from brain tissue.
Main Methods:
- Discontinuous Percoll gradient centrifugation for separating cellular components.
- Nitrogen cavitation to liberate mitochondria from synaptosomes.
- Further purification of mitochondria using Percoll gradient centrifugation.
Main Results:
- Effective separation of synaptosomes, myelin, and free nonsynaptic mitochondria.
- Successful isolation of synaptic mitochondria after nitrogen cavitation.
- Obtained mitochondria exhibit high respiratory coupling and rates.
Conclusions:
- The described protocol provides a reliable method for isolating highly pure and functional brain mitochondria.
- This technique is valuable for neuroscience research requiring intact mitochondria.
- The isolated mitochondria are suitable for studies on cellular respiration and mitochondrial function.
