Korean J Fertil Steril.  1997 Dec;24(3):347-353.

In Vitro/In Vivo Development after Thawing of Vitrified Mouse Blastocysts by Culture Condition and Embryo Transfer Method

• Kim MK
• Kim EY
• Yi BK
• Yoon SH
• Park SP
• Chung KS
• Lim JH

Abstract

SUMMARY: This study was to test whether in vitro/in vivo survival of vitrified mouse blastocysts was influenced by culture conditions and ET method. Mouse blastocysts were obtained from in vitro fertilization and cultured for 4 days in Ml6 medium, and they were vitrified in EFS40 which contained 40% ethlyene glycol, 18% Ficoll and 0.5 moi sucrose in PBS. In experiment 1, in vitro and in vivo survival rate of these embryos were evaluated in different culture condition after thawing. When thawed embryos were cultured in Ml6 medium as a control, m-CR1 medium contained 20 amino acids (2% BME amino acis and 1% MEM non-essential amino acids solution) and 4 mg/ml BSA and cumulus monolayer cell co-cultured condition in mCR1 medium (10% FBS), their in vitro survival at 24 hr after thawing was not affected by culture condition (75.6, 83.1, 82.4%). However, in vivo survival rates of implantation in m-CR1 medium (80.4%) were significantly higher than those of Ml6 medium (51.2%), coculture (57.1%) condition, although there was no difference in live fetuses rates on day 15 gestation (39.0, 49.0, 38.1%). In experiment II, the in vivo development potential of embryos by ET methods was examined. When blastocysts were transferred to the day 2, 3 pseudopregnant recipient without culture soon after thawing, no pregnant recipient was obtained on the day 2 pseudopregnancy, and 50% of pregnancy rates and 15.4% of live fetus rates were obtained on the day 3 pseudopregnant recipients. These results were significantly lower than those of transferred group (day 3 pseudopregnant recipients) after culture for 16 hr post thawing (73.5, 57.1%) (p<0.05). In experiment III, to elevate usability of delayed embryos in vitro/in vivo survival of vitrified embryos (day 4 early, day 5 early and expanding blastocyst) were examined. in vivo survival rates (live fetus, total implantation) were higher in day 4 early blastocysts (33.3, 66.7%) than in day 5 expanding blastocysts (29.0, 38.7%), although the highest in vitro survival rates were obtained in the day 5 expanding brastocysts (78.3%). Therefore, these results suggest that the in vitro/in vivo survival rates of vitrified embryos could be improve by the culture condition and ET method and that the in vivo development rates of delayed embryos were decreased with longer culture duration in vitro. It means that more effective cryopreservation was obtained in day 4 early blastocysts than in day 5 expanding blastocysts.

MeSH Terms

Amino Acids
Animals
Blastocyst*
Coculture Techniques
Cryopreservation
Embryo Transfer*
Embryonic Structures*
Fertilization in Vitro
Fetus
Ficoll
Mice*
Pregnancy
Pregnancy Rate
Pseudopregnancy
Sucrose
Survival Rate
Amino Acids
Ficoll
Sucrose