**2.4 Biological aspects of thawing**

While there are many risk factors associated with freezing of cells, thawing can also dramatically affect survival rates of spermatozoa. When frozen samples are returned to ambient temperature, a reversal of the freezing process takes place. Cells that were frozen by the slow method, are more vulnerable to rapid thawing, due to the fast influx of water into cells causing uncontrollable swelling and osmotic shock (Curry & Watson, 1994). If cells were frozen rapidly, intracellular ice crystals could re-crystallize and form larger crystals during a slow thaw. To minimize toxic effects, CPAs have to be promptly removed from the cell suspension by washing samples in isotonic solution. Therefore, the thawing process and CPAs removal technique utilized must take into account the original method that was used for freezing.
