Microvesicles were purified from the cell media by using a standardized protocol with slight modifications (7): conditioned cell media were collected and centrifuged for 10 min at 300 g for removing floating cells and debris. The resulting supernatants were further cleared through a 5-µm syringe-filter (Millex, Millipore), then ultracentrifuged at 10,000 g for 30 min to pellet MVs. The pellets obtained were resuspended in lysis buffer with protease inhibitor, PBS or fixative depending on the specific aim. As serum contains high levels of MVs, cells were cultured in Optimem or in DMEM supplemented with serum depleted of MVs by an overnight ultracentrifugation (at 110,000 g) as described in Shelke et al., (8).