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Solid-Surface Vitrification and In-Straw Dilution After Warming of In Vitro-Produced Bovine Embryos

Authors

  • P Rodriguez-Villamil,

    1. Instituto de Reproducción Animal Córdoba (IRAC), Córdoba, Argentina
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  • FL Ongaratto,

    1. Instituto de Reproducción Animal Córdoba (IRAC), Córdoba, Argentina
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  • M Fernandez Taranco,

    1. Instituto A.P. de Ciencias Básicas y Aplicadas, Area de Medicina Veterinaria, Universidad de Villa María, Villa María, Córdoba, Argentina
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  • GA Bó

    Corresponding author
    1. Instituto de Reproducción Animal Córdoba (IRAC), Córdoba, Argentina
    2. Instituto A.P. de Ciencias Básicas y Aplicadas, Area de Medicina Veterinaria, Universidad de Villa María, Villa María, Córdoba, Argentina
    • Author's address (for correspondence): Gabriel Bó, Instituto de Reproducción Animal de Córdoba, Córdoba, Argentina. E-mail: gabrielbo@iracbiogen.com.ar

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Contents

Three experiments were designed to test a solid-surface vitrification system for bovine in vitro-produced embryos and to develop a simple method of in-straw dilution after warming, which can be potentially used for direct transfer in the field. Experiment 1 evaluated embryo survival rates (i.e. re-expansion and hatching) after vitrification and warming in three different solutions: VS1 (20% ethylene glycol (EG) + 20% propanediol (PROH) + 0.25 m trehalose (Tr)), VS2 (20% EG + 1M Tr) or VS3 (30% EG + 0.75 m Tr). Re-expansion and hatching rates were higher (p < 0.05) for embryos vitrified in VS3 (72.2 ± 1.9 and 58.2 ± 0.8) than VS1 (64.4 ± 0.9 and 37.2 ± 2.5) or VS2 (68.5 ± 1.5 and 49.6 ± 1.0; p < 0.05). Experiment 2 was designed to compare two methods of vitrification: glass micropipettes or solid surface, using the VS1 or VS3 solutions. No significant differences were detected between the two methods; but re-expansion and hatching rates were higher (p < 0.05) with VS3 (73.5 ± 3.1 and 47.1 ± 2.1) than VS1 (63.3 ± 3.3 and 39.7 ± 2.8). In experiment 3, embryos were vitrified by solid surface in VS1 or VS3 solutions and cryoprotectants were diluted in-straw after warming in a TCM 199, 0.25 m sucrose solution or holding media. Survival rates of embryos vitrified in VS3 did not differ between those exposed to 0.25 m sucrose (74.7 ± 1.3 and 57.2 ± 2.2) or holding (77.3 ± 1.4 and 58.0 ± 2.5) medium after warming; however, survival rates of embryos vitrified in VS1 were higher (p < 0.05) in those exposed to 0.25 m sucrose (67.7 ± 2.3 and 47.0 ± 1.7) than holding medium (54.5 ± 1.0 and 27.7 ± 3.1). In conclusion, solid-surface vitrification using simplified EG-based solutions and in-straw dilution with holding media may be a practical alternative for cryopreservation and direct transfer of in vitro-produced bovine embryos.

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