Fig. 5

Polymer-based precipitation efficiently extracts functional exosomes from 7.5 × 106 cells. a SDS-page protein quantification showing that the greatest efficiency in terms of exosomal protein per cell plated was obtained when exosomes were extracted from differentiated myoblasts at a density of 33,400 cells.cm⁻². Differentiated myoblasts were plated at 14,147 (lane 1), 33,400 (lane 2), or 106,100 (lane 3) cells per cm². Right panel: representative SDS-page stained with Coomassie. Left panel: protein concentration measurements in secreted exosomes from cells plated at a different density. *, ***, P < 0.05 and P < 0.001, significantly different from 33,400 and 106,100 cells.cm². (n = 4, 3, 4 per condition). b Muscle exosomes were positive for CD63, CD81, Flotillin, HSPA8, and Alix. c mRNA was detectable with a clean profile from polymer-precipitated exosomes of 7.5 × 10⁶ differentiated myoblasts. No 18 s and 28 s RNA were detected, indicating that there were no RNA contaminants from dead cells. Inset panel: Representative electrophoresis obtain with Agilent 2100 Bioanalyzer for myotubes and exosome RNA extract. d Western blot showing that polymer precipitated exosomes from 7.5 × 10⁶ differentiated myoblasts can be used to pull down a specific subpopulation such as CD63 positive exosomes (+/-CD63 = with/without anti-CD63 antibody). e Polymer-precipitated exosomes (pre-stained with PKH26 following extraction; red channel) were capable of integrating into myotubes or into iPSC motor neuron cells