Utilizing tetraspanin-based isolation and LEGENDplexTM assay for MSC-derived exosome characterization 2724

Abstract Description Mesenchymal stem cells (MSCs) are highly recognized in regenerative medicine due to their multipotent differentiation and immunomodulatory properties. Recent studies suggest that the immunomodulatory effect of MSCs is mainly mediated through paracrine factors, with exosomes being critical players. This study focuses on the immunomodulatory effects of MSC-derived exosomes. MSCs were isolated from bone marrow and cultured in serum-free xeno-free media. Exosomes were extracted from the conditioned medium using a tetraspanin-based human exosome isolation kit, with purity confirmed via flow cytometry, western blot, and nanoparticle tracking analysis (NTA). The cargo of MSC-derived exosomes was characterized using LEGENDplexTM assay, a multiplex bead-based immunoassay system, revealing a diverse array of bioactive molecules crucial for tissue repair and immunomodulation. We investigated the immunomodulatory effects of these exosomes on CD3/CD28 activated T cells. Exosomes were depleted from the MSC conditioned media, and T cells were cultured in conditioned media with and without exosome-depletion. Flow cytometry and LEGENDplexTM analysis showed that MSC-derived exosomes significantly reduced T cell proliferation, altered cytokine profiles, and decreased inflammatory markers. These findings highlight the potential of MSC-derived exosomes as a cell-free therapeutic option in regenerative medicine. Funding Sources N/A Topic Categories Technological Innovations in Immunology (TECH)