Zetaview pmx 220

MiniSEC was used to isolate plasma-derived exosomes and was routinely performed as detailed in our previous publications [15 (link),51 (link)]. In short, plasma samples were centrifuged at 2000 g for 10 min, at 10,000 g for 30 min, and ultrafiltrated using a 0.22 µm filter to remove impurities. One milliliter of plasma was added to a miniSEC column and eluted with phosphate-buffered saline (PBS). The fourth fraction was collected and concentrated using Amicon Ultra columns (Merck Millipore, Burlington, MA, USA). Extracellular vesicles isolated from plasma were characterized according to the MISEV 2018 guidelines for the definition of extracellular vesicles [52 (link)]. Isolation of exosomes was confirmed by assessing particle size and count using nanoparticle tracking (ZetaView PMX-220, Particle Metrix), morphology by transmission electron microscopy, and origin from the endosomal compartment by Western Blot.

NTA was performed using the Zeta View PMX-220 (Particle Metrix, Inning, Bayern, Germany). Freshly isolated exosomes were diluted to a concentration of 1:100,000 in 1 mL PBS and loaded into the cell. Measurements were taken at 11 different positions throughout the cell. At each position, three cycles of records were taken. The instrument was operated at the following parameters: temperature of 25 °C, sensitivity of 85, and a shutter of 100. As a control, PBS and 100 nm polystyrene beads were used. The nanoparticle size ranges and concentrations were calculated using ZetaView 8.05.11 software.