Figure 1.

Carrier-based cell culture increases exosome yield. (A) Schematic overview of exosome production process. MenSC were expanded on 2D cell culture dishes and then seeded on BioNOC II. After 72 h in DMEM with 10% FBS, the medium was changed to serum-free DMEM for 72 h for exosome production. For purification, the supernatant was collected and processed in serial centrifugations. (B) Western Blot with 15 µg of cell lysate (Cells) and exosomes (Exo). To confirm the purity of the exosomes, positive exosomal markers CD63, Syntenin and CD9 and negative exosomal markers Vinculin (Vinc.) and Calreticulin (Calr.) and β-Actin (actin) were analyzed. (C) Scanning electron micrograph of purified exosomes, magnification 60,000x. (D) Size distribution of exosomes determined by nanosight showing that the highest abundance of particles was below 200 nm. (E) Hoechst-stained MenSC on BioNOC II carrier, showing a typical confluence for exosome production. (F) Yield of purified exosomes in PBS as Particles (part)/ml of initial cell culture supernatant (SN) are shown for 2D (N = 32) and 3D culture (N = 10). Analysis: unpaired t-test. Bar graphs show average values, error bars: SEM.