Cytofluorometric Assessment of Mitophagic Flux in Mammalian Cells and Tissues

Mitophagy is the selective autophagic degradation of mitochondria inside lysosomes and is the only mechanism capable of eliminating whole mitochondria that are either damaged or no longer required. Mitochondrial dysfunction is a common hallmark in many pathological conditions and has been implicated in cellular damage in aged tissues. The limited availability of mitophagy research methods underscores the need for more robust, quantitative, and objective tools in order to better understand this process. Here we describe a flow cytometry-based method using MitoTracker Deep Red for mitophagy assessment that can be applied to cells and tissues. Moreover, we demonstrate that when used in conjunction with lysosomal inhibitors, our method provides a novel means of assessing mitophagic flux, which is the most reliable indicator of whether mitochondria are truly delivered to lysosomes for degradation.