Open-pulled straw (OPS) vitrification of mouse hatched blastocysts.
This study was first employed to investigate the developmental potential of mouse hatched blastocyts (HBs) vitrified by a two-step open-pulled straw (OPS) method. HBs were obtained by culture of morulae in vitro. First, the embryos were placed in four cryprotectant solutions - that is, 10% ethylene glycol (EG), 10%E + 10%D (10% EG and 10% dimethyl sulphoxide (DMSO) in mPBS), EFS30 (30% EG, Ficoll, and sucrose) and EDFS30 (15% EG, 15% DMSO, Ficoll, and sucrose)--at 25 degrees C for 0.5 to 10 min, respectively, to determine their optimal survival after rapid dilution in 0.5 M sucrose. Secondly, based on the above best survival, the embryos were plunged into liquid nitrogen after first pretreatment in 10%E for 0.5 min and then 0.5 min equilibration in EFS30 (Group 1), or 10%E + 10%D and EDFS30 for 0.5 min, respectively (Group 2). When warming, three methods were used to dilute the cryoprotectants from the vitrified embryos. The embryos were assessed by the re-expansion of the blastocoel or development to term. The result showed that all the vitrified-warmed HBs got high in vitro survival rates (83.7% to 98.9%). The highest in vitro survival rates (87.8% in Group 1, 98.9% in Group 2) were obtained when the vitrified embryos were diluted first in 0.3 M sucrose for 5 min, then in 0.15 M sucrose for 2 min (method C). When the vitrified embryos diluted with method C were transferred, their survival rate in vivo (35.5% to 42.2% of the total) were similar to (P > 0.05) that of control (45.7%). These results demonstrate OPS method was highly efficient for the cryopreservation of mouse HBs.
Zhou
Chong
C
Laboratory of Animal Embryonic Biotechnology, College of Animal Science and Technology, China Agricultural University, Beijing, PR China.
Zhou
Guang-Bin
GB
Zhu
Shi-En
SE
Hou
Yun-Peng
YP
Jin
Fang
F
Zhao
Xue-Ming
XM
Hong
Qiong-Hua
QH
D000818:Animals; D001755:Blastocyst; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D003451:Cryoprotective Agents; D004121:Dimethyl Sulfoxide; D004624:Embryo Transfer / Q000662:veterinary; D019855:Ethylene Glycol; D005260:Female; D005362:Ficoll; D008297:Male; D051379:Mice; D009028:Morula / Q000166:cytology; D011247:Pregnancy; D013395:Sucrose
D003451:Cryoprotective Agents; D005362:Ficoll; D013395:Sucrose; D019855:Ethylene Glycol; D004121:Dimethyl Sulfoxide