Twin pregnancy after vitrification of 2-pronuclei human embryos

Fertil Steril. 2002 Feb;77(2):412-4. doi: 10.1016/s0015-0282(01)02992-2.


Objective: To report an ongoing twin pregnancy after transfer of embryos that were vitrified at the 2-pronuclei stage in a new vitrification solution.

Design: Case report.

Setting: A tertiary-care infertility clinic.

Patient(s): A 26-year-old infertile woman in whom two previous IVF implantations failed.

Intervention(s): Vitrification of 2-pronuclei embryos, in vitro culture for 48 hours, and transfer into the uterus.

Main outcome measure(s): Survival and cleavage after vitrification and achievement of clinical pregnancy.

Result(s): Six zygotes were vitrified by using a three-step protocol (4% ethylene glycol for 3 minutes, 20% ethylene glycol for 1 minute, and 38% ethylene glycol and 1.2 M trehalose for 0.5 minute). After 2 months of storage in a double-straw system in liquid nitrogen, two zygotes were warmed and cryoprotectants were removed by using a four-step protocol (1 M, 0.5 M, 0.25 M, and 0.125 M of trehalose). Two embryos were transferred after 48 hours of in vitro culture, cleaving to 5 and 6 cells. The resulting twin pregnancy was confirmed by ultrasonography at the sixth week.

Conclusion(s): Vitrification by using ethylene glycol and trehalose appears to be a safe, promising method for cryopreservation of human zygotes. Storage of vitrified zygotes in a double-straw system does not compromise their subsequent potential for survival and development.

Publication types

  • Case Reports

MeSH terms

  • Adult
  • Cryopreservation / methods
  • Cryoprotective Agents
  • Embryo Transfer*
  • Ethylene Glycol / pharmacology
  • Female
  • Fertilization in Vitro / methods*
  • Humans
  • Male
  • Pregnancy
  • Pregnancy, Multiple*
  • Trehalose / pharmacology
  • Twins


  • Cryoprotective Agents
  • Trehalose
  • Ethylene Glycol