Extracellular Vesicle Isolation Protocol

SCPs were isolated by differential centrifugation as adapted from the protocol for the isolation of extracellular vesicles (EVs) from [44 (link)]. The filtered homogenate was centrifuged twice at 300× g and 4 °C for 10 min in the centrifuge Centric 260R with rotor RA 6/50 (Domel, Železniki, Slovenia) by using 50 mL conical centrifuge tubes (ref. S.078.02.008.050, Isolab Laborgeräte GmbH, Germany). The supernatant of the second centrifugation was centrifuged twice at 2000× g and 4 °C for 10 min in the centrifuge Centric 400R with rotor RS4/100 (Domel, Železniki, Slovenia), using 15 mL conical centrifuge tubes (ref. S.078.02.001.050, Isolab Laborgeräte GmbH, Eschau, Germany). Then, the supernatant was centrifuged at 10,000× g or 50,000× g and 4 °C for 60 min in Beckman L8-70M ultracentrifuge, rotor SW55Ti (Beckman Coulter, Brea, CA, USA) using thin-wall polypropylene centrifuge tubes (ref. 326819, Beckman Coulter, Brea, CA, USA). The respective isolates were obtained by dissolving pellets in a small amount of solvent (e.g., 80 µL).