4.7. Isolation of SCPs from Plant and Microalgae Samples

SCPs were isolated by differential centrifugation, using a protocol widely used for the isolation of small extracellular vesicles [63]. Briefly, the cells were removed by low-speed centrifugation (300× g, 10 min, 4 °C, centrifuge Centric 260R with rotor RA 6/50 (Domel, Železniki, Slovenia)), using 50 mL conical centrifuge tubes (ref. S.078.02.008.050, Isolab Laborgeräte GmbH, Eschau, Germany); and 2000× g, 10 min, 4 °C (Centric 400R centrifuge with rotor RS4/100 (Domel, Železniki, Slovenia)), using 15 mL conical centrifuge tubes (ref. S.078.02.001.050, Isolab Laborgeräte GmbH, Eschau, Germany). Each step was repeated twice. Then, the cell-depleted medium was centrifuged twice at 10,000× g and 4 °C for 30 min (Beckman L8-70M ultracentrifuge, rotor SW55Ti (Beckman Coulter, Brea, CA, USA)), using thin-wall polypropylene centrifuge tubes (ref. 326819, Beckman Coulter, Brea, CA, USA) to remove larger cell debris. Finally, NPs were pelleted by ultracentrifugation at 118,000× g and 4 °C for 70 min in the same type of ultracentrifuge and ultracentrifuge tubes. The pellet was resuspended in 50 µL of the initial medium (PBS/ultraclean waste/marine water).