Principles of Ice-Free Cryopreservation by Vitrification

Vitrification is an alternative to cryopreservation by freezing that enables hydrated living cells to be cooled to cryogenic temperatures in the absence of ice. Vitrification simplifies and frequently improves cryopreservation because it eliminates mechanical injury from ice, eliminates the need to find optimal cooling and warming rates, eliminates the importance of differing optimal cooling and warming rates for cells in mixed cell type populations, eliminates the need to find a frequently imperfect compromise between solution effects injury and intracellular ice formation, and can enable chilling injury to be "outrun" by using rapid cooling without a risk of intracellular ice formation. On the other hand, vitrification requires much higher concentrations of cryoprotectants than cryopreservation by freezing, which introduces greater risks of both osmotic damage and cryoprotectant toxicity. Fortunately, a large number of remedies for the latter problem have been discovered over the past 35 years, and osmotic damage can in most cases be eliminated or adequately controlled by paying careful attention to cryoprotectant introduction and washout techniques. Vitrification therefore has the potential to enable the superior and convenient cryopreservation of a wide range of biological systems (including molecules, cells, tissues, organs, and even some whole organisms), and it is also increasingly recognized as a successful strategy for surviving harsh environmental conditions in nature. But the potential of vitrification is sometimes limited by an insufficient understanding of the complex physical and biological principles involved, and therefore a better understanding may not only help to improve present outcomes but may also point the way to new strategies that may be yet more successful in the future. This chapter accordingly describes the basic principles of vitrification and indicates the broad potential biological relevance of this alternative method of cryopreservation.