Ice forms at different rates during the cooling process.

As ice forms, water is removed from the extracellular environment and an osmotic imbalance occurs across the cell membrane leading to loss of water from the cell.

The increase in solute concentration can be detrimental to cell survival.

Too much water retention inside the cell cause damage to the cells and thus reduce the cell viability due to ice crystal formation and recrystallization during warming can occur.

The rate of cooling has a dramatic effect on over all cryopreservation process.

Rapid cooling minimizes the solute concentration and thus affects the uniform ice formion, but leads to more intracellular ice.

On the other hand, slow cooling, results in a greater loss of water from the cell and less internal ice, but increases the solution effects.

A cooling rate of 1°C per minute is generally preferred. Cryoprotective additives or chemicals, for example dimethylsulfoxide (DMSO) and glycerol can protect the cells during freezing and minimize the detrimental effects of increased solute concentration and ice crystal formation.

Cryopreservation media generally consists of a base medium, cryopreservative and a protein source.

The cryopreservative and protein protect the cells from the stress of the freeze-thaw process.

Additionally, it maintains frozen cells at the proper storage temperature and using an appropriate warming rate will minimize damage to frozen cells.