Evaluation of the impact of customized serum-free culture medium on the production of clinical-grade human umbilical cord mesenchymal stem cells: insights for future clinical applications

• Customized serum-free media (SFM) enhance hUC-MSC proliferation, activity, and stability compared to serum-containing media (SCM), while maintaining consistent morphology and surface markers. • Different SFM formulations yield distinct cellular behaviors (activity, proliferation, senescence, cell cycle), underscoring the need for tailored media design for specific clinical applications. • SFM-cultured hUC-MSCs retain normal karyotype after long-term culture, supporting their biosafety for clinical-grade production. • The study provides critical insights for industrial-scale production of clinical-grade MSCs, emphasizing cost-effectiveness, inter-batch consistency, and regulatory compliance.