In Vitro Budding and Floatation based Enrichment of Mitochondria-derived Vesicles for Proteomics from Rat Heart

Summary Mitochondria-derived vesicles (MDVs) are a novel class of vesicles whose biogenesis and trafficking are poorly characterized. Here, we describe the protocol to in vitro generate MDVs from mitochondria isolated from animal heart tissues. The protocol can be coupled with Transmission Electron Microscopy (TEM), proteomics or western blotting as final readouts. The protocol is optimized to improve the yield and purity of vesicles for downstream proteomics and facilitates selection of key regulators to be validated within cells. Graphical abstract The protocol has been adapted and further modified from (Soubannier et al., 2012).