Background

In prior research we demonstrated that freeze-dried stem cells stored for 3 years at room temperature were able to direct embryonic development following cloning. However, viability, as evaluated by membrane integrity of the cells after freeze-drying, was very low; and it was mainly the DNA integrity that was preserved. In a subsequent present study, we improved the cells’ viability and functionality after freeze-drying.

Methodology

We optimized the conditions of directional freezing, i.e. interface velocity and cell concentration, and we added the antioxidant EGCG to the freezing solution. The study was performed on mononuclear cells (MNCs) derived from human umbilical cord blood. After freeze-drying, we tested the viability, number of CD34+-presenting cells and ability of the rehydrated hematopoietic stem cells to differentiate into different blood cells in culture.

Principal findings

The viability of the MNCs after freeze-drying and rehydration with pure water was 88%–91%. The total number of CD34+-presenting cells and the number of colonies did not change significantly when evaluated before freezing, after freeze-thawing, and after freeze-drying (5.4×104±4.7, 3.49×104±6 and 6.31×104±12.27 cells, respectively, and 31±25.15, 47±45.8 and 23.44±13.3 colonies, respectively).

Conclusions

This was the first report of nucleated cells which were dried and then rehydrated with double-distilled water remaining viable, and of hematopoietic stem cells retaining their ability to differentiate into different blood cells.

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