Demarcating the membrane damage for the extraction of functional mitochondria.

Defective mitochondria have been linked to several critical human diseases such as neurodegenerative disorders, cancers and cardiovascular disease. However, the detailed characterization of mitochondria has remained relatively unexplored, largely due to the lack of effective extraction methods that may sufficiently retain the functionality of mitochondria, particularly when limited amount of sample is considered. In this study, we explore the possibility of modulating hydrodynamic stress through a cross-junction geometry at microscale to selectively disrupt the cellular membrane while mitochondrial membrane is secured. The operational conditions are empirically optimized to effectively shred the cell membranes while keeping mitochondria intact for the model mammalian cell lines, namely human embryonic kidney cells, mouse muscle cells and neuroblastoma cells. Unsurprisingly, the disruption of cell membranes with higher elastic moduli (neuroblastoma) requires elevated stress. This study also presents a comparative analysis of total protein yield and concentrations of extracted functional mitochondria with two commercially available mitochondria extraction approaches, the Dounce Homogenizer and the Qproteome