Chromatographic purification of small extracellular vesicles using an affinity column for phospholipid membranes

Abstract

Abstract Purpose: The use of small extracellular vesicles (sEVs) such as exosomes for therapeutic purposes has recently generated great interest. Developing an efficient, rapid, scalable, and reproducible sEV purification method is necessary. Therefore, this study aimed to investigate whether chromatography using an ExoPUA column, an affinity column for phospholipid membranes, could potentially meet this demand. Methods: We used the ExoPUA column connected to a fast-performance liquid chromatography system. Results: One-step chromatographic purification of sEVs from a culture supernatant using the ExoPUA protocol resulted in an 82-fold increase in purity with a yield of 38% of sEVs on average. Transmission electron microscopy and western blotting analysis indicated that purified sEVs were intact. Conclusion: The purification performance of the ExoPUA protocol yields superior results compared with that of the differential ultracentrifugation method, which is the most commonly used purification method for sEVs in the laboratory. Furthermore, sEVs were purified in the bind-elute mode and the ExoPUA column can be refreshed and sterilized with sodium hydroxide (NaOH), having more potential for multiple sEV purification in a scalable and industrial manner.