Quench cooling and ice crystal formation in biological tissues

The rates of cooling of tissue packages quenched in liquid nitrogen were investigated using microthermocouples. By assembling tissue packages from a standard 200-μm tissue slice a microthermocouple could be positioned at different depths within the package. Results showed that for a given mass of tissue the rates of cooling at different depths were the same. When the tissue mass was varied the rates of cooling at a fixed depth decreased with increasing tissue mass.The ice crystal formations produced when tissues are quenched in liquid nitrogen were investigated using freeze substitution. Assemblies of rabbit cornea of different thicknesses were quenched in liquid nitrogen and freeze substituted. The size of the ice crystal cavities produced during the quenching increased with increasing tissue mass, exhibiting a saturation size for the larger tissue masses. There was no obvious size distribution of the ice crystal cavities across the thickness of the corneas.The results suggest an “isotherm” model for the quenching conditions used in these experiments, there being small or negligible temperature gradients through the tissue which uniformly cools at a fixed rate.