Size Exclusion Chromatography for Exosome Isolation

EVs were purified from human serum samples using size exclusion chromatography on a drip with extracellular vesicle isolation by size exclusion chromatography on a EVSecond L70^® drip column (GL Sciences, Tokyo, Japan). The column was initially equilibrated with 700 μL of FBS twice, followed by three washing steps using 1500 μL of PBS. After washing, 100 μL of the collected human serum sample was loaded onto the column, followed by the collection of 12 consecutive fractions in 100 μL of PBS. The CD9 expression in these fractions was analyzed using western blotting, and CD9-positive fractions were recognized as the exosome-rich portion. The obtained EVs using this system were previously authenticated with TEM and western blotting, and the diameters of the particles in the EV fractions were analyzed with the Zetasizer nano ZSP [24 (link)].