Rs for instance cell passage and cell seeding density influence EV biogenesis and bioactivity has the prospective to improve therapeutic EV production. Right here, we investigate the influence of these parameters on MSC-derived EV production and vascularisation bioactivity. Strategies: Conditioned media was collected right after 24 h from MSCs seeded at unique densities (1E2, 5E2, 1E3, 1E4 cells/cm2) or passages (P2-P5). EVs have been isolated in the conditioned media through differential PLK2 drug centrifugation and quantified by nanoparticle tracking analysis (NTA) employing aScientific Plan ISEVNanosight LM10 and CD63 ExoELISA. Vascularisation bioactivity of isolated EVs was assessed in a wound healing assay. Final results: NTA and ExoELISA benefits indicated increased EV production prices per cell when MSCs have been seeded at lower initial densities, regardless of the cell passage. The typical fold lower in EVs production per cell amongst cells seeded at 1E2 cells/cm2 and 1E4 cells/cm2 for P2, P3, P4, and P5 was 100, 85, 110, and 50, respectively (n = five, p 0.01). Furthermore, several EV collection time points (12 and 24 h) in the identical cells enhanced total EV production far more than three fold compared to a single collection more than the exact same time period (24 h) (n = three, p 0.05). Seeding density had no affect on the vascularisation bioactivity of MSC EVs created as assessed by the wound-healing assay (n = three). In contrast, growing cell passage was correlated with diminished EV bioactivity (n = 3). Conclusion: These results suggest that higher EV production rates could be achieved by seeding cells at reduce initial seeding densities. Low cell passage quantity is essential to retaining MSC EV vascularisation bioactivity. The implications of these findings are that higher amounts of bioactive EVs is usually achieved making use of a reduced quantity of producer cells with enhanced frequency of collection. This might allow for significant reduction in cost of EV production and commence to inform the rational design and style of a large-scale biomanufacturing strategy for therapeutic EV production.eGFP-positive EVs. Cell proliferation kinetics inside bioreactors are monitored by glucose uptake, along with the production of EVs each under serum-supplemented and defined serum-free conditions is currently evaluated. The concentration and size distribution are measured by nanoparticle tracking analysis (NTA) and surface marker expression profiles and uptake kinetics in recipient cells of harvested EVs are analysed through flow Apical Sodium-Dependent Bile Acid Transporter Biological Activity cytometry. All parameters are when compared with classical 2D culture. Also, unique schedules for EV harvesting are compared as a way to optimise and standardise the production. Preliminary results and experiences using hollow fibre bioreactors for the large-scale production of EVs from distinct cell types will be presented here.PT02.Purifying and molecular profiling extracellular vesicles (EVs) from numerous biological specimens Abiodun Ogunjimi1 and Liang ZhangLunenfeld-Tanenbaum Investigation Institute; 2City University of Hong Kong, ChinaPT02.Evaluation and optimisation of a hollow fibre bioreactor system for standardisation of substantial scale production of extracellular vesicles Ulrika Felldin1, Giulia Corso1, Bernd Giebel1,two, Helmut Hanenberg3, Joel Z. Nordin1, Samir El-Andaloussi1,4 and AndrG gens1,1 Department of Laboratory Medicine, Karolinska Instiutet, Stockholm, Sweden; 2Institute for Transfusion Medicine, University Hospital Essen, University of Duisburg-Essen, Essen, Germany; 3Department of Pediatrics III, University C.