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Related Experiment Videos

Solid CO2 freezing of mouse embryos.

H Miyamoto, T Ishibashi

    Journal of Reproduction and Fertility
    |January 1, 1983
    PubMed
    Summary

    Rapidly freezing eight-cell mouse embryos using a two-step glycerol method resulted in high survival rates after thawing. This cryopreservation technique enables the development of normal young from frozen-thawed embryos.

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    Area of Science:

    • Reproductive biology
    • Cryobiology

    Background:

    • Cryopreservation of mammalian embryos is crucial for assisted reproduction and genetic resource preservation.
    • Optimizing freezing and thawing protocols is essential to maximize embryo survival rates.

    Purpose of the Study:

    • To investigate optimal conditions for rapid cryopreservation of eight-cell mouse embryos using a two-step cooling procedure.
    • To evaluate the viability and developmental potential of frozen-thawed mouse embryos.

    Main Methods:

    • Eight-cell mouse embryos were frozen using a two-step procedure involving crushed solid carbon dioxide (CO2) and liquid nitrogen, with glycerol as a cryoprotectant.
    • Key parameters optimized included glycerol addition, equilibration and holding times, and cooling/thawing rates.
    • Embryo survival was assessed after rapid and slow thawing, with subsequent transfer to recipient mice.

    Main Results:

    • Optimal survival was achieved with stepwise glycerol addition, no seeding, 1x 8 M-glycerol, 5-30 min equilibration, 10-30 min holding in solid CO2, and rapid thawing (~500°C/min).
    • A high survival rate of 62% was observed following rapid thawing, whereas slow thawing resulted in no survival.
    • Frozen-thawed embryos developed into normal young after transfer to recipient mice.

    Conclusions:

    • Rapid cryopreservation using a specific two-step cooling and rapid thawing protocol is effective for preserving eight-cell mouse embryos.
    • This method supports the developmental competence of mouse embryos, offering a viable option for assisted reproductive technologies.

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