Comparison between manual and automated methods for the isolation of mononuclear cells and mesenchymal stem cells using ficoll: efficacy and reproducibility

Ester Moñivas ^1* Concepción Aguayo ² Silvia de la Calle ² Paula Martínez ¹ Nieves Repollés ² Mercedes Zurita ²

• ¹ Instituto de Investigación Sanitaria Puerta de Hierro - Segovia de Arana, Hospital Universitario Puerta de Hierro Majadahonda, Majadahonda, Spain
• ² Puerta de Hierro University Hospital Majadahonda, Madrid, Madrid, Spain

Mesenchymal stem cells (MSCs), herein referred to as adherent cells, hold immense therapeutic promise in Advanced Therapy Medicinal Products (ATMPs) due to their multipotent nature, immunomodulatory properties, and anti-inflammatory effects.However, a significant challenge lies in obtaining sufficient quantities of MSCsadherent cells for therapeutic applications, necessitating ex-vivo culture of Mononuclear Cells (MNCs) isolated from source tissues like bone marrow. This study compares the efficacy of MNC isolation using manual and automated methods, specifically evaluating the Sepax system, and investigates whether the isolation method impacts MSCsadherent cell yield. Seventeen bone marrow samples were processed using both methods, with subsequent analysis of MNC and colony-forming unit (CFU) counts, MSCsadherent cell differentiation potential, and phenotypic characterization. While the Sepax system demonstrated slightly higher MNC yields, no significant differences were observed in CFU formation or MSCsadherent cell characteristics compared to manual isolation. These findings underscore the importance of critically evaluating isolation methods to ensure both efficiency and quality in therapeutic applications.