EVs purification
EVs purification was carried out from plasma samples by centrifugation
at 12,000g for 20 minutes at 10°C. Supernatants were transferred to
ultracentrifuge tubes (Beckman, Coulter), supplemented with PBS and
ultracentrifuged (XL-80, rotor 70Ti; Beckman, Coulter) twice at 100,000g
for 1 hour and 10 minutes at 10ºC. The pellet obtained was resuspended
in PBS and stored at 4ºC until its use in electron microscopy, in
vitro vascular permeability assays and proteomic analysis or -20ºC for
nanoparticle tracking analysis (NTA) and Western blot determination.