fig4

Figure 4. iMSC-EVs characterization. (A) Representative size exclusion chromatography (SEC) elution profile of iMSC-EV, positive for the EV marker CD63 and MSC marker CD90. Protein elution was measured at 280 nm absorbance by nanodrop and occurred later and separated from EV fractions; (B) Dot blot results for the detection of CD63 (top) and calnexin (bottom). 3D-derived samples from different weeks of EV-production of both bioreactors and 2D-derived samples from both iMSC lines were analyzed. The control (cellular lysate) is represented in duplicates; (C) Nanoparticle tracking analysis (NTA) showing the particle size distribution of 2D- and 3D-derived EVs. Data are normalized by mL of supernatant (left) or by million of producing cells (right); (D) iMSC-EV protein quantification by µBCA assay, showing the µg/mL of protein and (E) lipid quantification by Sulfo-phospho-vanillin (SPV) assay, showing the of µg/mL lipids, results normalized per mL of supernatant; (F) iMSC-EV protein quantification by µBCA assay, showing the µg/mL of protein and (G) lipid quantification by Sulfo-phospho-vanillin (SPV) assay, showing the of µg/mL lipids, results normalized per million producing cells. The samples represented are from two bioreactors (3D) and from five 2D-derived EV productions (2D); (H) Protein/lipid ratio for 3D and 2D samples. Bars represent means, and error bars represent SD. Comparison between groups was performed by Mann-Whitney Test. ns P > 0.05, *** P < 0.001, **** P < 0.0001.