2.3. EV characterization
The EV morphology was observed by
transmission electron microscopy (TEM). EVs were fixed with 2%
paraformaldehyde for 15 min; then, the mixture (about 8 μL) was dropped
onto carbon-coated copper grids and air-dried for 10 min before staining
with 1% uranyl acetate two times (6 min each). Images were obtained
using an FEI Tecnai BioTwin TEM (Thermo Fisher Scientific). To determine
the size distribution, dynamic light scattering (DLS), (Brookhaven
Instruments Corporation; Holtsville, NY) was utilized for early (0 day;
D0), middle (7 days; D7) and late (21-days; D21) time points of
osteogenic differentiation.