Exosomes are membrane-derived nanovesicles of 30 to 150 nm in diameter that are released by many cell types under both normal and pathological conditions. The influence of exosomes on cellular function has been linked to a wide range of physiological processes including: cell to cell communication, cancer metastasis, immunomodulatory activity, and propagation of infectious agents such as prions and retroviruses. Given this wide range of potential interactions, and considering the fact that these nanovesicles contain RNA, microRNA and proteins from their cells of origin, exosomes represent a burgeoning target for biomarker discovery.
The current gold standard for exosome isolation is differential ultracentrifugation. However, this method requires specific instrumentation, is lengthy, and labor intensive. Here we present a rapid alternative method for the selective fractionation of exosomes from cell culture media using an ultrafiltration device. Optimization of the protocol was aided through use of a mid infrared (MIR)-based spectroscopy platform that permits simultaneous monitoring of lysis conditions, protein quantitation, and analysis of total lipid content during exosome fractionation. The resulting exosome preps were analyzed using a NanoSight platform (to measure size distribution) and by flow cytometry of exosome-bead conjugates (for surface marker expression).