Culturing human umbilical cord blood

One of the main limiting factors for an increased use of human umbilical cord blood (UCB) in the adult allogeneic transplantation setting is the small number of progenitor cells that can be collected and infused. Ex vivo expansion of UCB might help overcome this limitation. Numerous culture methods have been applied to achieve cell number proliferation without inducing terminal differentiation. We here report on a comparison of UCB mononuclear cells (MNC) with CD34+ selected cells in a serum-free static culture system. Cell number proliferation of MNCs was inferior to CD34+ selected cells. MNCs, however, showed a substantial increase from 0.94% CD34+ cells on day 0 to 5.8% on day 7, whereas in the CD34+ selected samples the CD34+ cell content declined continuously from 62.2% on day 0 to 27.7% on day 7. To see whether an expansion of UCB also leads to co-expansion of contaminating maternal cells, male (XY) cord blood samples were investigated for maternal (XX) cells at day 0 and at several time points during culture conditions with interphase FISH analysis. We could not detect maternal cells in any of the 9 studied samples when cultures were started at day 0. Culturing did not expand previously undetected maternal cells into a range that could be seen with FISH technology. We then artificially contaminated male UCB with maternal mononuclear cells at concentrations of 5 and 15% at day 0. After 14 days maternal MNC were still detectable, but the percentage was reduced to 1.7% and 6%, respectively.