Successful cryopreservation of in vitro derived bovine blastocysts in microcapillary pipette tips

The open pulled straw has been used effectively to vitrify preimplantation embryos because of geometric features that allow rapid rates of temperature exchange. One possible inexpensive alternative to the open pulled straw are commercially-available microcapillary pipette tips commonly used for electrophoresis. The main purpose of this study was to compare the survival rates of in vitro produced blastocysts following vitrification in microcapillary pipette tips and open pulled straws. Two experiments were conducted to investigate the effect of type of carrier, age of expanded blastocyst, and addition of β-mercaptoethanol to post-warming culture medium on survival of vitrified in-vitro derived blastocysts. Expanded blastocysts (Day 7, 0900H and 1900H after insemination; insemination = Day 0 at 0900H) were vitrified while loaded randomly in groups of 4-10 into open pulled straws or pipette tips. Following warming, embryos were cultured in groups of up to 20 in 20 µl microdrops of modified KSOM or modified KSOM containing 100 µM β-mercaptoethanol at 38.5°C in 5% CO2 in air. Survival after warming was assessed as the percentage of vitrified embryos that re-expanded and the percent that hatched after 48 hrs culture. Post-warming survival rates were not affected by type of carrier or age of the expanded blastocyst (P>0.05). The proportions of embryos that re-expanded (55.5%) and hatched (25.7%) were higher (P<0.01) for those cultured with β-mercaptoethanol than for those cultured without (re-expansion: 42.4%; hatching: 12.6%). In conclusion, the microcapillary pipette tip represents an inexpensive alternative to the open pulled straw for cryopreservation.