Purification of human plasma exosomes for proteomics: optimization and application to detect changes in response to exercise

Extracellular vesicles (EVs) are released into blood from multiple organs and carry molecular cargo that facilitates inter-organ communication and an integrated response to physiological and pathological stimuli. Interrogation of the protein cargo of EVs is currently limited by the absence of optimal and reproducible approaches for purifying plasma EVs that are suitable for downstream proteomic analyses. We describe a size exclusion chromatography (SEC)-based method to purify EVs from platelet poor plasma (PPP) for proteomics profiling via high-resolution mass spectrometry (SEC-MS). The SEC-MS method identified more proteins with higher precision compared to several conventional EV isolation approaches. We applied the SEC-MS method to identify the unique proteomic signatures of EVs released from platelets, adipocytes, muscle cells, and hepatocytes, with the goal of identifying tissue-specific EV markers. Further, we applied the SEC-MS approach to evaluate the effects of a single bout of exercise on EV proteomic cargo in human plasma.