Fig. 1

Identification of SEC fractions for EV analysis via flow cytometry. A Schematic of experimental design. Twelve, 0.5 mL aliquots were collected after 1 mL of PFP was applied to SEC. B SEC fractions were analyzed via flow cytometry after FSC and SSC voltage was adjusted to reliably detect 1 μm beads (polystyrene microspheres) (top) and excluding mechanical noise observed in PBS alone (middle). Example of pooled SEC fractions seven, eight, and nine pooled from an individual donor (bottom). C The number of events within the size gate for each individual 0.5 ml SEC fractions was determine via flow cytometry. Fractions seven, eight, and nine (highlighted in red) contained the majority of detectable events and were pooled together for future analysis. D TEM analysis of pooled fractions from HC and RRMS patient. E Violin plots of EV diameter measured from TEM images for three HC and three RRMS patients. ns = nonsignificant determined by t-test. F Representative examples of NTA results of EV isolated from a HC and RRMS patient. G Western blot analysis of indicated proteins from purified EV lysates from three separate donors. HUVEC lysate was used as a positive control