Storage of unprocessed G-CSF-mobilized whole blood in a modified Leibovitz's L15 medium preserves clonogenic capacity for at least 7 days

Abstract

Summary:Autologous stem cell transplantation using unprocessed, G-CSF-mobilized whole blood (WB) is a simple, cost-reducing procedure and supports high-dose chemotherapy regimens not exceeding 72 h. Thereafter, clonogenic capacity rapidly decreases if routine anticoagulants are used for storage. In order to increase clinical applicability, we investigated the requirements for optimal preservation of unprocessed WB for 7 days. During storage at 22؇C in CPDA-1, a decrease in pH was noted, which was at least partially responsible for the low recovery of clonogenic cells. Subsequently, WB cells were stored in various cell culture media (RPMI 1640, ␣-MEM, X-VIVO15, CellGro SCGM and Leibovitz's L15 medium) containing either serum, serum-free substitutes or no additives. Leibovitz's L15 showed significantly better CFU-GM recoveries than the other media. Using a calcium-free modification of L15 medium (added 3:10 to WB), 94 ؎ 24% of CD34 ؉ cells, 41 ؎ 14% of BFU-E, 56 ؎ 17% CFU-GM and 90 ؎ 14% of LTC-IC were preserved during storage for 7 days at 22؇C. Storage at 4؇C was also feasible, but showed less optimal recoveries of 52 ؎ 29% (CD34), 32 ؎ 10% (BFU-E), 13 ؎ 7% (CFU-GM) and 58 ؎ 9% (LTC-IC). The expression of CD38, Thy-1, c-kit, AC133, L-selectin and CXCR4 on CD34-positive cells remained unchanged. In conclusion, a modified Leibovitz's L15 medium better meets the metabolic requirements of a high-density cell culture and allows safe storage of G-CSF mobilized WB for at least 7 days. The results encourage further exploration of WB transplants stored for 7 days for clinical use. Bone Marrow Transplantation (2001) 28, 145-155.

Granulocyte colony‐stimulating factor mobilized whole blood containing over 0·3 × 106/kg CD34+ cells is a sufficient graft in autologous transplantation for relapsed non‐Hodgkin's lymphoma

“…Although we did not compare LTC‐IC between PBSC and WB in this study, the WB results suggest that the frequency of LTC‐IC was not entirely different to apheresis samples (Boiret et al , 2001). Also, in a previous study, we found that primitive subsets in WB were comparable with normal PBSC collections (de Kreuk et al , 2001). Another explanation could be that the BAM regimen is not as myelotoxic as the BEAM regimen.…”

The analysis of viability for mammalian cells treated at different temperatures and its application in cell shipment

“…Strikingly, most of cells have been dead after stored at 1°C for 24 hours or 22°C for 72 hours (S3A Fig); suggesting that cryopreservation medium is more harmful to M2 cells than its own culture medium under hypothermia. It has been described that Leibovitz’s L15 medium can preserve blood cells and maintain their clonogenic capacity for 7 days [16]. Therefore, M2 and Hela cells were preserved in complete L15 medium under hypothermia, the result shows that L15 medium did not improve cell survival rate compared with their own culture mediums (S3B–S3E Fig, Figs 1D and 2E); however, mild hypothermia did enhance the viability for these cell lines compared with severe hypothermia.…”

“…The results from our study suggest that mammalian cells seem not suitable for incubation on ice or storage at 4 or 5°C, particularly for the cells that are sensitive to severe hypothermia or that need further culturing; thus, our finding extends the understanding on the effect of severe hypothemia on cell survival. IT has been described that hypothermic damage to cell can be improved by the utilization of conservation medium [16,19,20]. In this study the leibovitz’s L15 medium was used to store M2 and Hela cells under hypothermia, but no improvement had been observed for the viability.…”

Low‐oxygen and high‐carbon‐dioxide atmosphere improves the conservation of hematopoietic progenitors in hypothermia

“…Our results demonstrated that O 2 /CO 2 concentrations close to those in hematopoietic tissues (3% O 2 /6% CO 2 ) were beneficial for CFC preservation and CD34+ cells viability stored at 4°C in defined storage media. The preservation of more than 60% HPCs and approximately 70% Day 0 CD34+ cells over a 7‐day period provided some of the best results obtained among experiments with purified CD34+ cell populations, 6,19 whole apheresis product, 4 and diluted whole blood 20 . Moreover, it is approaching the high‐efficiency recovery of HCs after cryopreservation 21 …”