The laboratory of Prof. Naomi Nakagata at CARD-University of Kumamoto (Japan) has achieved a new milestone in mouse cryopreservation techniques. After boosting the efficiencies of mouse in-vitro-fertilization (IVF) from fresh, refrigerated and, particularly, frozen sperm from C57BL/6J mice, they have focused their attention in establishing a robust, reproducible but simple and efficient method for cryopreserving unfertilized mouse oocytes using vitrification, and their subsequent use for IVF. Their results have just been published in Cryobiology:
Applications of cryopreserved unfertilized mouse oocytes for in vitro fertilization.
Nakagata N, Takeo T, Fukumoto K, Kondo T, Haruguchi Y, Takeshita Y, Nakamuta Y, Matsunaga H, Tsuchiyama S, Ishizuka Y, Araki K.
Cryobiology. 2013 Oct;67(2):188-92.
The vitrification and thawing of unfertilized oocytes was one of the methods taught by the Nakagata-team, for the first time abroad, at the most recent CARD-CNB Mouse Sperm and Embryo Cryopreservation Course, held in Madrid two weeks ago. The efficient cryopreservation of unfertilized metaphase-II oocytes had remained a challenging procedure for investigators. Unfertilized oocytes are fragile germ cells that do not have nuclear membrane. However, these are most relevant cells and the ones to be used for somatic cell nuclear-transfer (SCNT), intracytoplasmic sperm injection (ICSI) and, in particular, for in vitro fertilization (IVF) techniques. Standard available vitrification methods failed to work, reproducibly, with unfertilized oocytes, whereas similar methods were routinely used with fertilized oocytes, for the efficient production of transgenic mice by pronuclear microinjection.
In 2013, a first report by Kohaya et al., described the generation of live offspring from vitrified C57BL/6J mouse oocytes for the first time. Now, Nakagata et al., show us in their most recent publication the efficient use of vitrified mouse unfertilized oocytes for their routine application to IVF techniques, in combination with fresh, refrigerated and frozen sperm, with high success. The resulting 2-cell embryos obained were transferred to recipients and also gave rise to pups at high rates. The popularization of these vitrification methods for unfertilized oocytes will enable a more efficient planification for IVF sessions. Using the stock of vitrified unfertilized oocytes, IVFs can be programmed daily, and should not be limited to the strict superovulation schedules that usually result in IVFs done only once (on Thursday) or twice (Tuesday and Thursday) per week.