Updated publication reference for PubMed record(s): 27068479. Polyethylene glycol (PEG) is commonly used to isolate virus particles from culture medium. This project adapted and optimized the virus procedure for harvesting extracellular vesicles (of similar biophysical properties to viruses) from media. An 8% PEG solution harvested the purest vesicles. Followed by a wash step (ultracentrifugation of concentrated vesicles in saline), this method produced preparations comparable or superior to commonly used methods in terms of purity, abundance, and cost. It is also highly adaptable, allowing the procedure to be used for enriching vesicles from a variety of biological fluids. As proof of principle, extracellular vesicle protein was successfully harvested from HeLa cell medium using the described PEG-based harvest method and analyzed by mass spectrophotometry.