Viability of CD34+ Cells in Cryopreserved Cord Blood

Purpose: On performing umbilical cord blood (UCB) transplantation, faster engraftment may lead better clinical outcome. Because transplanted viable cell count in UCB is related to the engraftment, we examined cryopreserved UCB cells with several methods after thawing.

Methods: Viability of cryopreserved cells were examined with trypan blue, DNA contents analysis, caspase-3 activation test, intracellular esterase activity and Annexin-V/PI staining.

Results: A total of 60 samples were used in this study. After thawing, 89% of the total MNCs and 84% of CD34+ cells were viable as identified by trypan blue exclusion assay. In the CD34+ cell population, the cell death rate was found to be 47 % by Annexin-V/PI staining and less than 5 % by DNA contents analysis. Caspase-3 activity failed to document apoptosis. The intracellular esterase activity test also showed a cell death rate of about 10–20 % at 2, 4, and 6 hours after thawing.

Conclusion: Viable cells in UCB should be measured by several compensatory techniques rather than a single method. Discordance among Annexin-V/PI staining versus trypan blue exclusion, DNA contents analysis, and the caspase-3 activation test or intracellular esterase activity should be clarified in order to apply these techniques for actual cord blood transplantation.