Effects of hold time after extracellular ice formation on intracellular freezing of mouse oocytes

MII mouse oocytes in 1 and 1.5 M ethylene glycol(EG)/phosphate buffered saline have been subjected to rapid freezing at 50 °C/min to −70 °C. When this rapid freezing is preceded by a variable hold time of 0-3 min after the initial extracellular ice formation (EIF), the duration of the hold time has a substantial effect on the temperature at which the oocytes subsequently undergo intracellular ice formation (IIF). For example, in 1 M EG, the IIF temperatures are −23.7 and −39.2 °C with 0 and 2 min hold times; in 1.5 M EG, the corresponding IIF temperatures are −29.1 and −40.8 °C.