Effects of cryobiological variables on the survival of skin using a defined murine model

Skin from an inbred strain of hairless mouse was used as a homogeneous model tissue for studies of skin cryopreservation. Tetrazolium reductase enzyme activity was used to assess tissue viability. Hepes-buffered 199 tissue culture medium was confirmed to be a suitable basal medium, to which cryoprotectants were added. Addition of serum to the cryoprotective cocktail had no beneficial effect. Three cryoprotectants, dimethyl sulfoxide, ethanediol, and glycerol were evaluated. There was no evidence of specific toxicity attributable to the cryoprotective agents during the permeation period; however, short permeation times at low temperature were associated with maximum skin viability. Following freezing and thawing, higher viabilities were obtained when using a slow (-1 degree C min-1) or medium (-60 degree C min-1) rather than a fast (immersion in liquid nitrogen) cooling rate. Dimethyl sulfoxide was a marginally better cryoprotectant overall, although this difference was not statistically significant.