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Comparison of two-step vitrification versus controlled freezing on survival of in vitro produced cattle embryos

(1994) THERIOGENOLOGY. 42(8). p.1389-1397
Author
Organization
Abstract
Grade 1 in vitro produced bovine day 7 embryos at the compact morula-early blastocyst, blastocyst and expanded blastocyst stage were selected for cryopreservation. From 7 replicates, 572 embryos were randomly divided into two soups. One soup was cryopreserved by controlled freezing after 20 minutes equilibration in 10% v/v glycerol and slow cooling (0.3 degrees C / minute) from -7 degrees C to -30 degrees C. The other group was vitrified after 3 minutes exposure to 20% v/v ethylene glycol and 30 to 45 seconds exposure to a vitrification solution consisting of 40% v/v ethylene glycol, 18% w/v ficoll and 10.26% w/v sucrose. Embryos from both soups were thawed in a water bath at 20+/-1 degrees C. Frozen-thawed embryos were diluted in three successive solutions consisting of 10.26% w/v sucrose and 6.6%, 3.3% and 0% glycerol, respectively, during 5 minutes for each seep. Vitrified-warmed embryos were diluted in a 8.58 w/v sucrose solution during 5 minutes, All diluted embryos were cultured in Menezo-B2 medium supplemented with bovine oviduct epithelial cells. The survival rate of all developmental stages of embryos was significantly higher after vitrification than after controlled freezing (P<0.001). It was 39.4% and 11.2% for compact morulae-early blastocysts, 81.9% and 34.9% for blastocysts and 96.7% and 72.3% for expanded blastocysts, respectively. The overall hatching rate following the two cryopreservation methods was not significantly different. However, the hatching rate of vitrified expanded blastocysts was significantly higher than the hatching rate of frozen expanded blastocysts (71.38 and 43.3%, respectively, P<0.001). It is concluded that the chilling sensitivity of embryos is significantly reduced by vitrification. A high proportion of blastocysts and expanded blastocysts can be successfully cryopreserved with a two-step vitrification procedure.
Keywords
IN VITRO EMBRYO, BOVINE, VITRIFICATION, CONTROLLED FREEZING, BOVINE EMBRYOS, INVITRO, CRYOPRESERVATION, VIABILITY

Citation

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Chicago
Mahmoudzadeh, AR, Ann Van Soom, Maria Ysebaert, and Aart de Kruif. 1994. “Comparison of Two-step Vitrification Versus Controlled Freezing on Survival of in Vitro Produced Cattle Embryos.” Theriogenology 42 (8): 1389–1397.
APA
Mahmoudzadeh, A., Van Soom, A., Ysebaert, M., & de Kruif, A. (1994). Comparison of two-step vitrification versus controlled freezing on survival of in vitro produced cattle embryos. THERIOGENOLOGY, 42(8), 1389–1397.
Vancouver
1.
Mahmoudzadeh A, Van Soom A, Ysebaert M, de Kruif A. Comparison of two-step vitrification versus controlled freezing on survival of in vitro produced cattle embryos. THERIOGENOLOGY. 1994;42(8):1389–97.
MLA
Mahmoudzadeh, AR, Ann Van Soom, Maria Ysebaert, et al. “Comparison of Two-step Vitrification Versus Controlled Freezing on Survival of in Vitro Produced Cattle Embryos.” THERIOGENOLOGY 42.8 (1994): 1389–1397. Print.
@article{200225,
  abstract     = {Grade 1 in vitro produced bovine day 7 embryos at the compact morula-early blastocyst, blastocyst and expanded blastocyst stage were selected for cryopreservation. From 7 replicates, 572 embryos were randomly divided into two soups. One soup was cryopreserved by controlled freezing after 20 minutes equilibration in 10\% v/v glycerol and slow cooling (0.3 degrees C / minute) from -7 degrees C to -30 degrees C. The other group was vitrified after 3 minutes exposure to 20\% v/v ethylene glycol and 30 to 45 seconds exposure to a vitrification solution consisting of 40\% v/v ethylene glycol, 18\% w/v ficoll and 10.26\% w/v sucrose. Embryos from both soups were thawed in a water bath at 20+/-1 degrees C. Frozen-thawed embryos were diluted in three successive solutions consisting of 10.26\% w/v sucrose and 6.6\%, 3.3\% and 0\% glycerol, respectively, during 5 minutes for each seep. Vitrified-warmed embryos were diluted in a 8.58 w/v sucrose solution during 5 minutes, All diluted embryos were cultured in Menezo-B2 medium supplemented with bovine oviduct epithelial cells. The survival rate of all developmental stages of embryos was significantly higher after vitrification than after controlled freezing (P{\textlangle}0.001). It was 39.4\% and 11.2\% for compact morulae-early blastocysts, 81.9\% and 34.9\% for blastocysts and 96.7\% and 72.3\% for expanded blastocysts, respectively. The overall hatching rate following the two cryopreservation methods was not significantly different. However, the hatching rate of vitrified expanded blastocysts was significantly higher than the hatching rate of frozen expanded blastocysts (71.38 and 43.3\%, respectively, P{\textlangle}0.001). It is concluded that the chilling sensitivity of embryos is significantly reduced by vitrification. A high proportion of blastocysts and expanded blastocysts can be successfully cryopreserved with a two-step vitrification procedure.},
  author       = {Mahmoudzadeh, AR and Van Soom, Ann and Ysebaert, Maria and de Kruif, Aart},
  issn         = {0093-691X},
  journal      = {THERIOGENOLOGY},
  keyword      = {IN VITRO EMBRYO,BOVINE,VITRIFICATION,CONTROLLED FREEZING,BOVINE EMBRYOS,INVITRO,CRYOPRESERVATION,VIABILITY},
  language     = {eng},
  number       = {8},
  pages        = {1389--1397},
  title        = {Comparison of two-step vitrification versus controlled freezing on survival of in vitro produced cattle embryos},
  url          = {http://dx.doi.org/10.1016/0093-691X(94)90259-L},
  volume       = {42},
  year         = {1994},
}

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