Fig. 3

The level of total proteins in the sEVs is reduced in Ubl3 knockout mice. a The cell lysate (CL) and pellets from the conditioned medium of MDA-MB-231 cells transfected with 3xFlag-UBL3 vectors were blotted with various antibodies. b The presence of UBL3 and its mutants in sEVs. c Electron microscopic analyses of purified sEVs by negative staining. Scale bars, 100 nm. d Upper left panel, protein staining for the sEVs from the serum in WT and Ubl3 KO mice. Lower panels, purified serum sEVs were blotted with anti-tubulin and CD9 antibodies with βME. Right panel, relative intensity of total proteins in serum sEVs. n = 9 pairs. e Total RNA levels in the serum sEVs. WT, n = 5; KO, n = 5. n.s., p > 0.05 by Mann–Whitney test. a,b, βME + condition: Flag, Flotillin-1, GP96, Actinin-4, Calreticulin, GAPDH, and Alix antibodies. βME- condition: CD63, and CD9 antibodies