Comparison of stem cell viability of bone marrow cryopreserved by two different methods

Area of Science:

• Hematology
• Cryobiology
• Cell Biology

Background:

• Cryopreservation is crucial for preserving hematopoietic stem cells.
• Classical methods involve cell separation and liquid nitrogen storage, which can be complex and lead to cell loss.

Purpose of the Study:

• To evaluate a modified cryopreservation technique for murine bone marrow cells.
• To compare the efficacy of a new method using unfractionated cells with a mixed protectant at -80°C against classical methods.

Main Methods:

• Murine bone marrow cells were cryopreserved using two methods: classical (separated cells, 10% DMSO, -196°C) and modified (unfractionated cells, 5% DMSO + 6% HES, -80°C).
• Cell viability (trypan blue exclusion) and recovery of colony-forming unit-granulocyte-macrophage (CFU-GM) and colony-forming unit-spleen (CFU-S) were assessed after 1, 4, 8, and 12 weeks of storage.

Main Results:

• The modified DMSO/HES method at -80°C resulted in high cell viability (87%) and recovery of CFU-GM (75%) and CFU-S (74%) after 3 months.
• These results were comparable to the classical method using fractionated cells at -196°C.
• Minimal cell loss (approx. 25% reduction in stem cells) occurred during cryopreservation itself, with no further deepening over time.

Conclusions:

• The cryopreservation of unfractionated murine bone marrow cells using a 5% DMSO and 6% HES mixture at -80°C is an effective technique.
• This simplified method offers high stem cell recovery and avoids complex cell-separation procedures, making it suitable for clinical applications.