Valenti 2014 Anal Biochem

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Valenti D, de Bari L, De Filippis B, Ricceri L, Vacca RA (2014) Preservation of mitochondrial functional integrity in mitochondria isolated from small cryopreserved mouse brain areas. Anal Biochem 444:25-31.

» PMID: 24018341

Valenti D, de Bari L, De Filippis B, Ricceri L, Vacca RA (2014) Anal Biochem

Abstract: Studies of mitochondrial bioenergetics in brain pathophysiology are often precluded by the need to isolate mitochondria immediately after tissue dissection from a large number of brain biopsies for comparative studies. Here we present a procedure of cryopreservation of small brain areas from which mitochondrial enriched fractions (crude mitochondria) with high oxidative phosphorylation efficiency can be isolated. Small mouse brain areas were frozen and stored in a solution containing glycerol as cryoprotectant. Crude mitochondria were isolated by differential centrifugation from both cryopreserved and freshly explanted brain samples and were compared with respect to their ability to generate membrane potential and produce ATP. Intactness of outer and inner mitochondrial membranes was verified by polarographic ascorbate and cytochrome c tests and spectrophotometric assay of citrate synthase activity. Preservation of structural integrity and oxidative phosphorylation efficiency was successfully obtained in crude mitochondria isolated from different areas of cryopreserved mouse brain samples. Long-term cryopreservation of small brain areas from which intact and phosphorylating mitochondria can be isolated for the study of mitochondrial bioenergetics will significantly expand the study of mitochondrial defects in neurological pathologies, allowing large comparative studies and favoring interlaboratory and interdisciplinary analyses.

Keywords: Brain mitochondria; Cryopreservation; Mitochondria isolation; Mitochondrial oxidative phosphorylation

Labels: MiParea: Respiration, Instruments;methods, mt-Membrane 

Stress:Cryopreservation, Ischemia-reperfusion  Organism: Mouse  Tissue;cell: Nervous system  Preparation: Isolated mitochondria  Enzyme: Marker enzyme 

Coupling state: OXPHOS, ET  Pathway: N, S, ROX