Fig. 5
a Histograms of side scatter area (SSC-A) of control PBS, showing the gating choice above electronic noise and below impurities, and SK-OV-3 EVs, showing the additional events both within and above the noise limit. c Histogram of SK-OV-3 EV size converted to diameter using FCMPASS calibration. d Dilution series of SK-OV-3 EVs illustrating constant median fluorescent intensity (MFI, violet circles) as events increased with concentration (gray squares), confirming that detected events are single particles in the concentration range tested. d Total number of fluorescent particles above background for labeled EVs versus antibody control. e Percent of total fluorescent EVs identified by each tetraspanin. Error bars represent ± one s.d. * p < 0.05, ** p < 0.01, *** p < 0.001 by Student’s t-test versus Ab control for (d) and ANOVA for (e). N = 3 experimental replicates