title: Assessment of cord blood hematopoietic cell parameters before and after cryopreservation.
abstract: BACKGROUND
  The testing of cord blood (CB) progenitor and stem cell units for transplantation suitability involves enumeration of total nucleated cells before freezing. CD34+ cell counts may also be a means of determining suitability. Studies have been conducted to evaluate how specific storage conditions influence cell counts.
  
  
  METHODS
  CB units were processed by hydroxyethyl starch volume reduction. Cryopreserved-thawed samples were diluted 1:3 without washing. CD34+ cells were measured with three commercially available assay methods. In specific studies, apoptosis-indicating reagents were included. CB units were analyzed for nucleated cells, aldehyde dehydrogenase-containing cells, and progenitor colonies.
  
  
  RESULTS
  CD34+ cell levels and nucleated cells were retained during storage in test tubes at 1 to 6 degrees C for 3 days. Cryopreserved-thawed samples showed a reduction in CD34+ cells relative to prefreeze levels with the largest decrease with the Stem-Kit (Beckman Coulter) restricted gating procedure. Prefreeze samples contained minimal numbers of presumed apoptotic cells detected with 7-aminoactinomycin D or SYTO16, but after cryopreservation-thawing there was an increase. Nucleated cell levels determined with a hematology analyzer or flow cytometry were reduced after thawing. Cryopreservation-thawing reduced the percentage of CD34+ cells positive for the presence of aldehyde dehydrogenase and the number of progenitor colonies. These differences were significant.
  
  
  CONCLUSIONS
  These studies indicate that CD34+ cell counts were maintained when CB samples were stored at 1 to 6 degrees C in test tubes for 3 days. Cryopreservation-thawing resulted in changes in a number of parameters including the percentage of CD34+ cells that were aldehyde dehydrogenase(+) and the number of 7-aminoactinomycin D(+) cells and SYTO16(low) cells.
authors.affiliation: American Red Cross, Jerome H. Holland Laboratory for the Biomedical Sciences, Rockville, Maryland 20855, USA. kurtzj@usa.redcross.org
authors.forename: James
authors.initials: J
authors.lastname: Kurtz
authors.forename: Shalini
authors.initials: S
authors.lastname: Seetharaman
authors.forename: Nicholas
authors.initials: N
authors.lastname: Greco
authors.forename: Gary
authors.initials: G
authors.lastname: Moroff
mesh_terms: D018952:Antigens, CD34 / Q000378:metabolism; D017209:Apoptosis; D015415:Biomarkers; D002467:Cell Nucleus; D003114:Colony-Forming Units Assay; D015925:Cryopreservation / Q000379:methods*; D005312:Fetal Blood / Q000166:cytology / Q000378:metabolism; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism; D006801:Humans
chemical_list: D018952:Antigens, CD34; D015415:Biomarkers