S. Murayama et al., SUCCESSFUL FREEZING OF UNFERTILIZED MOUSE OOCYTES AND EFFECT OF COCULTURES IN OVIDUCTS ON DEVELOPMENT OF IN-VITRO FERTILIZED EMBRYOS AFTER THAWING, Journal of assisted reproduction and genetics, 11(3), 1994, pp. 156-161
Purpose: To establish a freeze-thawing method for unfertilized oocytes
with a high success rate, we examined several conditions for freeze-t
hawing. The effects of EDTA and cocultures in oviducts on the developm
ent of embryos fertilized in vitro after thawing were also studied. Re
sults: In the first experiment, unfertilized oocytes that were frozen
in 1.5 M dimethylsulfoxide (DMSO) supplemented with 0.2 M sucrose by a
slow freeze-thawing method showed the best results (fertilization rat
e, 71.9%; blastocyst rate per frozen oocyte, 18.8%). The proportion of
embryos that developed to blastocysts was significantly higher when D
MSO was added at 4-degrees-C than at room temperature (39.4 vs 19.4%;
P < 0.01). The addition of EDTA (10 muM) to the culture medium did not
promote embryo development after fertilization in vitro. However, the
rate of development of in vitro fertilized embryos to blastocysts aft
er thawing was significantly higher when the embryos were cultured in
oviducts in vitro than the rates in control cultures and those culture
d with EDTA (blastocyst rate from fertilized oocytes, 71.4 vs 51.0 and
52.8%, respectively; P<0.01). Conclusion: Unfertilized mouse oocytes
can be cryopreserved successfully by a slow freeze-thawing method with
the addition of 1.5 M DMSO and 0.2 M sucrose at low temperatures, and
coculture with oviducts enhances the development of embryos that are
fertilized in vitro after thawing.