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Updated: May 9, 2026
Morphometric Protocol for the Objective Assessment of Blastocyst Behavior During Vitrification and Warming Steps
Published on: February 28, 2019
Effect of vitrification on number of inner cell mass in mouse blastocysts in conventional straw, closed pulled straw,
1Physiology Research Center, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran. ghasemsaki@yahoo.com
Objective:
To compare the effect of using open and closed carriers on count of inner cell mass in vitrified mouse blastocyst after warming.
Methods:
The experimental study was conducted at Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran, from April to September 2010. Forty female NMRI (Naval Medical Research Institute, USA) mice were injected with pregnant mare's serum gonadotropin and human chorionic gonadotropin in order to induce superovulation. Following the latter injection, two or three females were caged with the same-breed male mice. The presence of vaginal plug was examined the following morning. To collect blastocyst embryos, the pregnant females were sacrificed by cervical dislocation at 88-90 hours after the injection and dissected. Blastocysts were collected in phosphate-buffered saline and allocated to four groups: vitrification in conventional straw, closed pulled straw, open pulled straw and cryoloop. The vitrification solution was ethylene glycol, Ficol and sucrose (EFS) 20% and 40%. After storage for 1 month in liquid nitrogen, the blastocysts were thawed in 0.5 M sucrose then cultured in M16 medium. After 6 hours of culture, the number of expanded blastocysts was recorded and stained by double-dye technique. After staining, the number of total cell and inner cell mass was calculated.
Results:
The re-expansion rate of blastocysts in the cryoloop group (n = 90; 78.26%) was significantly higher (p < 0.05) than open pulled straw (n = 83; 69.16%), closed pulled straw (n = 68; 54.83%) and conventional straws (n = 63; 51.21%) groups. Significant differences (p < 0.05) in the number of inner cell mass in blastocysts vitrified in open pulled straws, closed straws and cryoloop with blastocysts cryopreserved in conventional straws.
Conclusion:
The re-expansion rate and total cell number of mouse blastocysts vitrified using open system had a better result compared with the closed system. The value of cryoloop and open pulled straws as carriers in vitrification of blastocysts may also merit more investigation.
