The viability of cryopreserved PBPC depends on the DMSO concentration and the concentration of nucleated cells in the graft

Background DMSO is widely used as a cryoprotectant for PBPC. It is desirable to reduce the amount of DMSO without jeopardizing the quality of the stem cell product. The present study was undertaken to investigate whether recovery and survival of CD34(+) cells would be significantly altered when PBPC used for autologous transplantations were cryopreserved with four different DMSO concentrations. Methods Apheresis samples of PBPC from 20 consecutive patients were mixed in parallel with 2%, 4%, 5% and 10% DMSO, frozen with identical cell concentrations at a controlled rate, and stored in liquid nitrogen for 6-8 weeks. PBPC samples from 11 consecutive patients were also cryopreserved with two different cell concentrations (150 and 300x10(6) nucleated cells/mL) to investigate the effect of increasing the cell concentrations while decreasing the DMSO concentration. The flow cytometric absolute count method, based on ISHAGE guidelines, was used to measure the absolute count of total and viable CD34(+) cells in the post-thaw samples. Results PBPC cryopreserved at 150x10(6) cells/mL with 2% DMSO yielded significantly inferior CD34(+) cell recovery (P<0.001) and survival (P<0.001) compared with cryopreservation with 4% and 5% DMSO. This was also observed when comparing higher cell concentrations. However, a reduced cell survival (P=0.02) was observed when the nucleated cell concentration was increased from 150 to 300x10(6) cells/mL in samples cryopreserved with 5% DMSO. Discussion We conclude that 5% DMSO may be the optimal dose for cryopreserving PBPC as long as the cells have not been concentrated at much more than 200x10(6) nucleated cells/mL.