Highly effective strategy for isolation of mononuclear cells from frozen cord blood.

ABSTRACT

BACKGROUND AIMS: Cord blood mononuclear cells (CBMCs) comprise a variety of single-nucleated cells found in the cord blood, mainly consisting of monocytes and lymphocytes. They also include a smaller proportion of other cell types, such as hematopoietic stem and progenitor cells (HSPCs) and mesenchymal stromal cells (MSCs). CBMCs are vital for acquiring HSPCs, MSCs, and other immune cells, like natural killer cells. These cells are essential for supporting subsequent research and clinical applications. Although automated equipment for CBMC enrichment has shown promise, the high cost of these machines and the expense of disposable consumables limit their routine use. Furthermore, limited information is available on manual strategies for isolating CBMCs from cryopreserved cord blood. Therefore, we aimed to optimize the dilution buffer and refine the isolation procedure for CBMCs. METHODS: We enhanced the CBMC recovery rate from cryopreserved cord blood using an optimized dilution buffer and a modified isolation procedure. RESULTS: We achieved average recovery rates of 42.4 % and 54.3 % for CBMCs and CD34+ cells, respectively. Notably, all reagents used in the isolation procedure were of GMP-grade or pharmaceutical preparations, underscoring the potential clinical benefits of our strategy. DISCUSSION: We devised an optimized protocol suitable for routine research and clinical applications for enhanced recovery of CBMCs from cryopreserved cord blood units using an optimized dilution buffer and a modified isolation procedure.