Direct ex vivo expansion of hematopoietic stem cells from umbilical cord blood on membranes

We have developed a direct ex vivo hematopoietic stem cell (HSC) expansion method involving filtration of umbilical cord blood (UCB) through polyurethane foaming membranes. Most of the red blood cells and mononuclear cells flow through the membranes, but HSCs remain on the membranes. The use of this method reduces the working time to less than 30 min before culture of HSCs, while conventional purification of HSCs from UCB that have been purified by the Ficoll–Paque procedure, followed by magnetic-activated cell sorting, results in loss of cells (i.e., a yield less than 15% of HSCs), damages HSCs, and takes a long time to perform (e.g., 5–8 h). Following filtration of the UCB and washing of the membranes, the membranes can be placed into culture medium where the HSCs can be expanded in a three-dimensional (3D) environment ex vivo, such as the bone marrow niche. Direct ex vivo expansion of HSCs from UCB resulted in an increased number of cells (i.e., 6.6- to 45.7-fold), and the expanded cell populations showed good hematopoietic ability in colony-forming unit assays.