fig3

Figure 3. Quantification of EVs isolated by ultracentrifugation, size exclusion chromatography, or EE affinity column. EVs were isolated from 4, 2, and 1 mL of acellular BAL by SEC, UC, and EE. Preparations were then analyzed for (A) particle yield by Nanosight (n = 3 subjects); (B) protein concentration (n = 3 subjects); (C) assessment of EV transmembrane proteins tetraspanin and syntenin-1 by western blotting; (D) Quantitation of western blot bands (n = 3 subjects); (E) Further confirmation of EV isolation was performed by detection of the intracellular EV protein TSG101. EVs: Extracellular vesicles; BAL: bronchoalveolar lavage; SEC: size exclusion chromatography; UC: ultracentrifugation; EE: exoEasy; CD: cluster of differentiation; Syn-1: syntenin-1; TSG101: tumor susceptibility gene 101.