Figure 11. Purified EMVs isolated from infected progenitor cells contained infectious virus.

(A) Supernatants from differentiated C2C12 cell cultures infected with eGFP-CVB3 were collected at day 3 PI and purified by Isopycnic gradient centrifugation. Buoyant density of infectious, membrane-associated EMVs released by C2C12 cells were observed within a range of 1.04–1.10 g cm⁻³. In contrast, a separate peak of infectious virus was observed at a density expected for non-enveloped virions (1.22 g cm⁻³). (B) Shed microvesicles isolated from differentiated C2C12 cells infected with eGFP-CVB3 were collected at day 3 PI and purified utilizing the Exoquick-TC polymer-based exosome precipitation kit. Purified EMVs were resuspended in 1× DMEM and inspected by fluorescence microscopy for viral protein (eGFP) and size distribution using the Length Interactive Measurement feature in AxioVision software. (C) Purified EMVs were added to HeLa cell cultures, and virus protein expression was followed by fluorescence microscopy over 24 hours PI.