Vitrification of human embryos based on the assessment of suitable conditions for 8-cell mouse embryos.
Experiments were conducted to find a suitable cryoprotectant and suitable procedure for vitrification of 8-cell mouse embryos. The method was then applied clinically to the cryopreservation of human embryos in our assisted reproduction programme. Mouse embryos were vitrified with 30 or 40% 1,2-propanediol (PROH), dimethylsulphoxide (DMSO), ethylene glycol, glycerol, or acetamide, each diluted with a solution containing 30% Ficoll plus 0.5 M sucrose. Embryos were exposed to the solutions for 0.5 or 2 min at 20 or 25 degrees C, cooled in liquid nitrogen and warmed rapidly. Embryo survival was assessed by in-vitro development. In PROH-, DMSO- and acetamide-based solutions, higher survival rates (29-82%) were obtained with less permeating conditions, suggesting that these cryoprotectants are considerably toxic. In glycerol- and ethylene glycol-based solutions, however, higher survival rates (74 and 92% respectively) were obtained with more permeating conditions, suggesting that these cryoprotectants are less toxic. Human embryos on days 2-3 were vitrified in an ethylene glycol-based solution (EFS40). Survival, assessed by the morphology, was higher in 4-cell embryos on day 2 and 8-cell embryos on day 3 than in 2-3-cell embryos on day 2 or 2-7-cell embryos on day 3. From 18 transfers, one ended with the delivery of healthy twin babies.
Mukaida
T
T
Hiroshima HART Clinic, Japan.
Wada
S
S
Takahashi
K
K
Pedro
P B
PB
An
T Z
TZ
Kasai
M
M
D000818:Animals; D002452:Cell Count; D002970:Cleavage Stage, Ovum / Q000166:cytology* / Q000187:drug effects; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000633:toxicity; D004121:Dimethyl Sulfoxide; D004624:Embryo Transfer; D004622:Embryo, Mammalian / Q000187:drug effects; D019855:Ethylene Glycol; D005069:Evaluation Studies as Topic; D005260:Female; D005307:Fertilization in Vitro; D005990:Glycerol; D006801:Humans; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D011247:Pregnancy; D019946:Propylene Glycol; D012099:Reproductive Techniques
D003451:Cryoprotective Agents; D019946:Propylene Glycol; D019855:Ethylene Glycol; D005990:Glycerol; D004121:Dimethyl Sulfoxide