In-straw Cryoprotectant Dilution for Bovine Embryos Vitrified Using Cryotop

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  • INABA Yasushi
    National Livestock Breeding Center, Fukushima 961-8511, Japan National Institute of Livestock and Grassland Science, Ibaraki 305-0901, Japan
  • AIKAWA Yoshio
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • HIRAI Tomokazu
    National Livestock Breeding Center Tokachi Station, Hokkaido 080-0572, Japan
  • HASHIYADA Yutaka
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • YAMANOUCHI Tadayuki
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • MISUMI Koji
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • OHTAKE Masaki
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • SOMFAI Tamas
    National Livestock Breeding Center, Fukushima 961-8511, Japan National Institute of Livestock and Grassland Science, Ibaraki 305-0901, Japan
  • KOBAYASHI Shuji
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • SAITO Norio
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • MATOBA Satoko
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • KONISHI Kazuyuki
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • IMAI Kei
    National Livestock Breeding Center, Fukushima 961-8511, Japan

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説明

The aim of this study was to develop an in-straw dilution method suitable for 1-step bovine embryo transfer of vitrified embryos using the Cryotop vitrification-straw dilution (CVSD) method. The development of embryos vitrified using the CVSD method was compared with those of embryos cryopreserved using in-straw vitrification-dilution (ISVD) and conventional slow freezing, outside dilution of straw (SFODS) methods. In Experiment 1, in vitro-produced (IVP) embryos cryopreserved using the CVSD method were diluted, warmed and exposed to the dilution solution at various times. When vitrified IVP embryos were exposed to the dilution solution for 30 min after warming, the rates of embryos developing to the hatched blastocyst stage after 72 h of culture (62.0-72.5%) were significantly lower (P<0.05) than those of embryos exposed to the solution for 5 and 10 min (82.4-94.3%), irrespective of supplementation with 0.3 M sucrose in the dilution solution. In Experiment 2, the rate of embryos developing to the hatching blastocyst stage after 48 h of culture in IVP embryos cryopreserved using the SFODS method (75.0%) was significantly (P<0.05) lower than those of embryos cryopreserved using the CVSD and ISVD methods (93.2 and 97.3%, respectively). In Experiment 3, when in vivo-produced embryos that had been cryopreserved using the CVSD, ISVD and SFODS methods and fresh embryos were transferred to recipient animals, no significant differences were observed in the conception and delivery rates among groups. In Experiment 4, when IVP embryos derived from oocytes collected by ovum pick-up that had been cryopreserved using the CVSD and ISVD methods and fresh embryos were transferred to recipient animals, no significant differences were observed in the conception rates among groups. Our results indicate that this simplified regimen of warming and diluting Cryotop-vitrified embryos may enable 1-step bovine embryo transfer without the requirement of a microscope or other laboratory equipment.<br>

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