Cryopreservation of animal cells

A wide range of cells of biological, medical and agricultural importance can be cryopreserved at -196 degrees C for many years in a stable state. The survival after freezing and thawing of different types of cell varies markedly, and depends on the ability of the cells to withstand a variety of stresses imposed by the physical and physico-chemical changes occurring in the bathing medium during cooling to and warming from the storage temperature. In most instances, cells will survive freezing only when a cryoprotectant (an additive such as glycerol or dimethyl sulphoxide that can protect cells against freezing injury) is included in the bathing medium: moreover, survival is dependent on the rates at which cells are cooled and warmed. Each cell type has an optimum cooling rate at which survival is maximal. This optimum cooling rate is determined to a large extent by the water permeability of the cell; but it is also influenced by the presence of cyroprotectants, and the warming rate. Therefore, when devising a method of cryopreservation for a particular cell type, the most important variables are the rates of cooling and warming, and the type and concentration of cryoprotectant. In general, as the concentration of cryoprotectant is increased, cell survival improves cryoprotectants can have detrimental toxic and osmotic effects on cells. Therefore, the way in which cryoprotectants are added to the bathing medium before freezing and, in particular, the procedure used to return the cells to a medium without cryoprotectants after thawing, can be crucial to the survival of the cells. Vitrification could be a way to avoid the mechanisms of injury associated with the formation of ice. When an aqueous solution contains a sufficiently high concentration of solute (e.g. greater than 40% w/w of the cyroprotectant propane-1,2-diol), the large increase in viscosity that occurs during cooling inhibits ice formation, and the solution eventually becomes an amorphous, glassy solid. There is no formation of a crystalline solid, nor is there a progressive rise in solute concentration, both of which occur as a result of ice formation during cooling. Vitrification could, therefore, prove to be a useful alternative method of cryopreservation, particularly for organized tissues and organs.(ABSTRACT TRUNCATED AT 400 WORDS)