Effect of AAGP™ on Maintaining Stem Cell Number and Function in Mouse Neural Stem Cells During 4°C Refrigeration for 2 Days

Hussein, August 2007

Introduction

Previous studies into the affect of AAGP™ on maintaining cell number and function in low density mouse neural stem cells during Cryopreservation showed AAGP™ to maintain cell function and increase cell survival 3 fold, in comparison to control cells not exposed to AAGP™. Therefore, the aim of this experiment was to investigate if the apparent protective properties of AAGP™ were affective when cells were exposed to refrigeration (4°C) for 2 days.

Method

Low density cell cultures of mouse neural embryonic stem cells were counted, exposed to AAGP™ or kept as a control and preserved via refrigeration at 4°C for 48 hours. Post-refrigeration the cell cultures were allowed to grow for 2 days. Population cell number was calculated on days 0, 1 and 2 post refrigeration using Trypan blue (4 counts per population).

The effectiveness of AAGP™ in maintaining cell number as a percentage of cells prior to Cryopreservation was measured in mouse embryonic neural stem cells in the same way. We used both high and low densities of mouse embryonic stem cells in one cell line and a high density of mouse neural stem cells in another cell line.

Results

As can be seen from the graph AAGP™ demonstrated a 5 fold increase in cell survival at day 1 and, probably due to cell proliferation, a 8-12 fold increase in cell survival at day 2. The functional competence of the stem cells was found to be 96% via OCT4 staining and flow cytometry.

Conclusion

Thus, it can be concluded that AAGP™'s apparent protection against cell death in low-density mouse neural stem cells is affective when cells are exposed to 4°C refrigeration as well as Cryopreservation. In addition, this mechanism of cell protection does not appear to affect the functionality of the stem cells. It could be said that this makes AAGP™ an attractive additive in the preservation media of stem cells for transplants where only a low density of cells can be extracted. These experiments will need to be completed in other cell lines.