title: Isolation of functionally active and highly purified neuronal mitochondria from human cortex.
abstract: BACKGROUND
  Functional and structural properties of mitochondria are highly tissue and cell dependent, but isolation of highly purified human neuronal mitochondria is not currently available.
  
  
  METHODS
  We developed and validated a procedure to isolate purified neuronal mitochondria from brain tissue. The method combines Percoll gradient centrifugation to obtain synaptosomal fraction with nitrogen cavitation mediated synaptosome disruption and extraction of mitochondria using anti mitochondrial outer membrane protein antibodies conjugated to magnetic beads. The final products of isolation are non-synaptosomal mitochondria, which are a mixture of mitochondria isolated from different brain cells (i.e. neurons, astrocytes, oligodendrocytes, microglia) and synaptic mitochondria, which are of neuronal origin. This method is well suited for preparing functional mitochondria from human cortex tissue that is surgically extracted.
  
  
  RESULTS
  The procedure produces mitochondria with minimal cytoplasmic contaminations that are functionally active based on measurements of mitochondrial respiration as well as mitochondrial protein import. The procedure requires approximately four hours for the isolation of human neuronal mitochondria and can also be used to isolate mitochondria from mouse/rat/monkey brains.
  
  
  CONCLUSIONS
  This method will allow researchers to study highly enriched neuronal mitochondria without the confounding effect of cellular and organelle contaminants.
authors.affiliation: Neuroapoptosis and Translational Therapeutics Laboratory, Department of Neurological Surgery, University of Pittsburgh, Pittsburgh, PA, United States.
authors.forename: Nicolas K
authors.initials: NK
authors.lastname: Khattar
authors.affiliation: Neuroapoptosis and Translational Therapeutics Laboratory, Department of Neurological Surgery, University of Pittsburgh, Pittsburgh, PA, United States.
authors.forename: Svitlana
authors.initials: S
authors.lastname: Yablonska
authors.affiliation: Neuroapoptosis and Translational Therapeutics Laboratory, Department of Neurological Surgery, University of Pittsburgh, Pittsburgh, PA, United States.
authors.forename: Sergei V
authors.initials: SV
authors.lastname: Baranov
authors.affiliation: Neuroapoptosis and Translational Therapeutics Laboratory, Department of Neurological Surgery, University of Pittsburgh, Pittsburgh, PA, United States.
authors.forename: Oxana V
authors.initials: OV
authors.lastname: Baranova
authors.affiliation: Neuroapoptosis and Translational Therapeutics Laboratory, Department of Neurological Surgery, University of Pittsburgh, Pittsburgh, PA, United States.
authors.forename: Eric S
authors.initials: ES
authors.lastname: Kretz
authors.affiliation: Neuroapoptosis and Translational Therapeutics Laboratory, Department of Neurological Surgery, University of Pittsburgh, Pittsburgh, PA, United States.
authors.forename: Timothy M
authors.initials: TM
authors.lastname: Larkin
authors.affiliation: Neuroapoptosis and Translational Therapeutics Laboratory, Department of Neurological Surgery, University of Pittsburgh, Pittsburgh, PA, United States.
authors.forename: Diane L
authors.initials: DL
authors.lastname: Carlisle
authors.affiliation: Brain Modulation Laboratory, Department of Neurological Surgery, University of Pittsburgh, Pittsburgh, PA, United States; University of Pittsburgh Medical Center, Pittsburgh, PA, United States.
authors.forename: R Mark
authors.initials: RM
authors.lastname: Richardson
authors.affiliation: Neuroapoptosis and Translational Therapeutics Laboratory, Department of Neurological Surgery, University of Pittsburgh, Pittsburgh, PA, United States; University of Pittsburgh Medical Center, Pittsburgh, PA, United States. Electronic address: friedlanderr@upmc.edu.
authors.forename: Robert M
authors.initials: RM
authors.lastname: Friedlander
mesh_terms: D000906:Antibodies / Q000378:metabolism; D002458:Cell Fractionation; D002540:Cerebral Cortex / Q000166:cytology*; D006680:HLA Antigens / Q000378:metabolism; D006801:Humans; D053078:Membrane Potential, Mitochondrial / Q000502:physiology; D008928:Mitochondria / Q000378:metabolism / Q000502:physiology*; D033681:Mitochondrial Membrane Transport Proteins / Q000276:immunology; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins / Q000378:metabolism; D009474:Neurons / Q000648:ultrastructure*; D013574:Synaptosomes / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D000906:Antibodies; D006680:HLA Antigens; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins; C489971:TOMM22 protein, human
keywords: Differential centrifugation; Functional; Human cortex; Isolation; Mitochondria; Neurons; Non-synaptosomal mitochondria; Percoll; Purified; Synaptosome