Fig 7. Single miRNAs are very scarce in EVs.
Three different preparations of EVs were obtained from conditioned medium of LCLs infected with wild-type EBV. EVs were purified by differential centrifugation (‘miniUC pellet’; Fig 2A), by size exclusion chromatography (SEC, fractions 7–9 were combined and analyzed; Fig 3A), or by iodixanol (Optiprep) density gradient centrifugation (fractions 2 and 3 were combined and analyzed; Fig 1D). Concentrations of EVs were determined by NTA. RNA was isolated from the three preparations (resuspended miniUC pellet, SEC and density gradient fractions), and absolute copy numbers of four different miRNAs were determined using a TaqMan stem loop RT-qPCR. For each miRNA, a standard curve of a synthetic RNA oligonucleotide (mimicking the miRNA) was established to determine absolute miRNA copy numbers in the preparations. (A) The graph shows the miRNA copy number per EV of three viral miRNAs and the human miRNA hsa-miR-16. (B) The graph indicates the miRNA copy numbers contained in 106 EVs (y-axis) as measured by RT-qPCR and NTA, respectively, versus the miRNA copy number contained in 106 cells (x-axis).