Optimising cryopreservation protocols for haematopoietic progenitor cells: a methodological approach for umbilical cord blood.
Current cryopreservation protocols for haematopoietic cells have developed largely empirically and there is no consensus on an optimal method of preservation. These protocols, though providing sufficient cells to permit engraftment, can lead to cell loss of the order of 50 percent. In the context of umbilical cord blood such losses are unacceptable. Whilst an empirical approach can provide an acceptable level of recovery, the cryopreservation process can only be optimised by adopting a methodological approach. This paper provides an overview of just such an approach as illustrated by a study on CD34 cells from umbilical cord blood. It involves firstly the determination of membrane permeability parameters that can then be used to model safe addition and elution protocols for the chosen cryoprotectant, in this case dimethyl sulphoxide. This in turn permits cryoprotectant toxicity to be evaluated free from the confounding effect of osmotic damage caused by inappropriate addition and elution protocols. Finally, non-toxic concentrations of cryoprotectant may be investigated in a cooling rate study to provide an optimal cryopreservation protocol. Using the model, the effect on CD34 cells of current addition and elution protocols was also examined.
Hunt
Charles J
CJ
UK Stem Cell Bank, National Institute for Biological Standards and Control, UK. chunt@nibsc.ac.uk
Pegg
David E
DE
Armitage
Susan E
SE
D018952:Antigens, CD34; D001793:Blood Preservation / Q000379:methods; D002463:Cell Membrane Permeability; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000009:adverse effects / Q000494:pharmacology; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D005312:Fetal Blood / Q000166:cytology*; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000473:pathology / Q000502:physiology*; D007700:Kinetics; D009994:Osmolar Concentration; D009995:Osmosis
D018952:Antigens, CD34; D003451:Cryoprotective Agents; D004121:Dimethyl Sulfoxide