Effects of cryopreservation on the viability and fertilizability of unfertilized hamster oocytes.

To examine the effects of cryopreservation on unfertilized oocytes, oocytes were collected from hamsters after injection of 25 IU of pregnant mare serum gonadotropin and 50 IU of human chorionic gonadotropin. Oocytes were placed in French straws containing Hepes-buffered Tyrode's medium and 2 mol/L of dimethyl sulfoxide at room temperature (method 1) or at 0 degree C (method 2). They were cooled to -6 degrees C at a rate of -2 degrees C/min (method 1) or at -0.5 degree C/min (method 2). The freezing rate was -0.3 degree C/min to a temperature of -80 degrees C, and then it was plunged to -196 degrees C. After thawing 1 to 21 days later at 8 degrees C/min, 52% (method 1) and 64% (method 2) of the oocytes showed normal morphologic features, 87% (method 1) and 89% (method 2) were viable by trypan blue staining, and 56% (method 1) and 59% (method 2) were viable by fluorescein diacetate staining. Comparable values for freshly collected oocytes were 95%, 100%, and 100%, respectively. Frozen-thawed oocytes were fertilized in vitro at a level of 26% (17/66) compared with 76% (51/67) for freshly collected controls. When the penetration rate of human spermatozoa was examined, frozen-thawed oocytes were penetrated at about one third of the fresh controls (14% or 18/129 compared to 42% in method 1 and 22% or 21/95 compared to 58% in method 2). The ability of oocytes to tolerate storage at -196 degrees C could be important in regard to various genetic studies, supply for hamster egg penetration assay, and cryopreservation of human oocytes.