Viability and Colony Forming Capacity of Hematopoietic Stem Cells after Cryopreservation

Jang, Sung Jun; Park, Euk; Park, Set Byul; Jung, Su Kyung; Kim, Heung Sik; Kang, Chin Moo

Abstract

PURPOSE: Cryopreservation of hematopoietic stem cells is one of the essential components in autologous and peripheral blood stem cell transplantation. Cryopreservation of hematopoietic stem cell, the conventional method involves controlled-rate freezing by a programmed freezer in medium that contains 10% dimethyl sulfoxide (DMSO) as cryoprotectant, followed by storage in liquid nitrogen freezer. We compared the differences between different methods of cryopreservation and cryoprotectants on viability and colony forming capacity of hematopoietic stem cells.
METHODS
Mononuclear cells separated using Ficoll-Hypaque from cord blood, peripheral blood and bone marrow were frozen with programmed freezer at 196degrees C or placed in a 70degrees C freezer without programmed freezer in both 10% and 20% DMSO. We measured cell viability using trypan blue dye exclusion method and colony forming capacity with methyl cellulose media at 7, 30 and 90 days after thawing.
RESULTS
Cell viability of cord blood, peripheral blood and bone marrow was higher in the groups with programmed freezer compared with rapid freezing and storing in a 70degrees C freezer. Also as the storage time passed, the decrease in viability of hematopoietic cells was much less in the groups of controlled-rate freezing by a programmed freezer. The number of colony in cord blood and bone marrow was higher with programmed freezer and that of peripheral blood was higher with rapid freezing and storage in a 70degrees C freezer. Comparing the differences between different concentraions of DMSO, cell viability was similar or slightly higher in 20% DMSO groups than 10% DMSO groups, but the number of colony was higher in 10% DMSO groups.
CONCLUSION
These results suggested that conventional cryopreservation method using programmed freezer with 10% DMSO was more effective in the cryopreservation of hematopoietic stem cells.