Abstract
The effect of cooling rate on the post-thaw viability of cells in cryopreserved artificial tissue was studied. Human fibroblasts were three-dimensionally cultured for 2 days in collagen sponge (φ20x1mm) as an extracellular matrix to imitate biological tissue (artificial tissue). Different cell densities for the artificial tissue were used, from 10^5 to 10^7 cells/cm^3. Four artificial tissues were first stacked in a test chamber, then frozen at a cooling rate from 0.3 to 50℃/min in a solution of Dulbecco's Modified Eagle Medium, 20% Fetal Bovine Serum, and 10% dimethylsulfoxide, then kept frozen at -196℃ for 2 hours, and finally thawed. Collagen matrix of artificial tissue was resolved by using collagenase. Post-thaw viability of fibroblasts was evaluated by using a trypan blue exclusion assay. Results show that with increasing cell density, the post-thaw viability decreased. Moreover, when the cell density was high, dehydration was obstructed, and thus the optimal cooling rate became slower.
