Fig 2: Multiple Myeloma bone marrow EVs have elevated CD59 and CD147 and decreased CD55 and CD9 levels compared to peripheral blood EV, whereas CD38 and PD-L1 are similar across all EV samples. 1.25 × 107 EVs/test from 23 matched BM and PB EVs from patients progressing on DARA were bound to the surface of 4 µm aldehyde/sulfate latex beads and stained with antibodies for; CD9, CD63, CD81, CD147 CD38, PD-L1 and the complement inhibitory proteins CD55 and CD59 for 30 min on ice. The samples were then resuspended in 200 µL PBS and flow cytometry analysis was performed on a CytoFLEX LX Flow Cytometer. Gating of EV-decorated 4 µm in diameter beads was performed based on FCS/SSC parameters, so that unbound EVs or possible antibody aggregates are excluded from the analysis.