EV-containing pellets were processed as described in our previous papers.12 (link),36 (link) Briefly, pelleted EVs were fixed with 4% paraformaldehyde at room temperature for 1 h, rinsed by PBS, and post-fixed in 1% osmium tetroxide (OsO4) for 20 min. After rinsing with distilled water, pellets were dehydrated by a series of increasing ethanol concentrations, including block staining with 1% uranyl-acetate in 50% ethanol for 30 min, finally embedded in Taab 812 (Taab, Aldermaston, UK). Following polymerization at 60°C for 12 h, 50–60 nm ultrathin sections were cut using a Leica UCT ultramicrotome (Leica Microsystems, Wetzlar, Germany, UK) and examined using a Hitachi 7100 transmission electron microscope (TEM) (Hitachi Ltd.,Tokio, Japan).
Electron micrographs were made by Veleta 2k x 2k Megapixel side-mounted TEM CCD camera (Olympus, Tokio, Japan). Contrast/brightness of electron micrographs was edited by Adobe Photoshop CS4 (Adobe Systems Incorporated, San Jose, CA, USA).
Electron micrographs were made by Veleta 2k x 2k Megapixel side-mounted TEM CCD camera (Olympus, Tokio, Japan). Contrast/brightness of electron micrographs was edited by Adobe Photoshop CS4 (Adobe Systems Incorporated, San Jose, CA, USA).