Figure 1. Cryopreservation marginally affects MSC viability and metabolic activity.

(A) MSC harvested from culture or thawed directly out of cryostorage were assayed for double strand DNA breaks by TUNEL staining. MSC were analyzed immediately after thawing or after 1 hour of storage on wet ice by flow cytometry and fluorescence imaging. The percent of positive and negative stained cells is reported in the upper right and left corners of each plot respectively. Cells were fixed, stained with PI (red) and FITC-dUTP (green). Double positive cells were considered dead. (Scale Bar = 100 μm). (B) Viability of MSC plated after thawing was compared to donor and passage matched MSC from fresh cultures 24, 48, and 72 hours after thawing. Cells were stained with Hoechst 33342 and PI and imaged with a fluorescence microscope. Cells double positive for Hoechst 33342 and PI were considered dead. (One-way ANOVA with Sidak correction for multiple comparisons, p < 0.05 considered significant, n = 5). (C) The metabolic activity of MSC after cryopreservation was compared to donor and passage matched MSC from continuous cultures using XTT (mean ± SD, One-way ANOVA with Sidak correction for multiple comparisons, p < 0.05 considered significant, n = 6). All experiments performed with MSC from donors 8002L and 7083 at passages P3–P5.