KMID : 1025520060480060797
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Journal of Animal Science and Technology 2006 Volume.48 No. 6 p.797 ~ p.804
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Effect of Sucrose Concentration on Survival After Frozen-thawed of Bovine IVF Blastocysts in Ethylene Glycol Based Freezing Medium for Slow-Cooling
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Cho S. R.
Kim H- J. Choi C. Y. Jin Hyun-Ju Son D. S. Choi Sun-Ho
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Abstract
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The present study was undertaken to investigate the post-thawed survivability of bovine embryo depending on different dose of ethylene glycol and sucrose. Ovaries were collected at local slaughterhouse and the cumulus-oocyte-complexes aspirated from ovaries were in vitro matured, fertilized and cultured at 39¡ÆC in an atmosphere of 5% CO2 incubator. For conventional slow-freezing, d 7 or 8 expanded blastocysts were collected. Embryos were equilibrated in 1.5 M and 1.8 M ethylene glycol(EG) with 0.1 M and 0.3 M sucrose in Dulbecco¡¯s phosphate-buffered saline(D-PBS) supplemented with 0.5% bovine serum albumin. Embryos were then loaded individually into 0.25ml-straw and placed directly into cooling chamber of programmable freezer precooled to 7¡ÆC, after 2 min, the straw was seeded, maintained at 7¡ÆC for 8 min, and then cooled to 35¡ÆC at 0.3¡ÆC/min, plunged and stored in liquid nitrogen for at least 3 days. For thawing, the straw containing embryos were warmed in air for 10 sec and exposed to 37¡ÆC water for 20 sec. Straws were then removed from 37¡ÆC water. Rates of blastocyst survive and hatching were evaluated at 24 to 72 h post-warming. No difference of the survivability was shown between 1.5 M and 1.8 M EG (71 and 70%, respectively). Addition of 0.1 M sucrose to 1.5 M and 1.8 M ethylene glycol in the freezing solution did not differ significantly embryo survival (74 and 77%, respectively), whereas survival rates was higher(89%) in freezing solution contained 0.3M sucrose to 1.8M EG compared with 0.3M sucrose to 1.5M EG group(71%). However, there was no difference in the overall total cell number between the two groups (122¡¾1.8 vs 131¡¾1.4, respectively). In conclusion, the results suggest that 0.3 M sucrose in 1.8 M EG may be optimal condition for freezing and thawing methods with in vitro produced embryos and may be applied to on-farm conditions for embryo transfer.
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KEYWORD
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Bovine, In vitro, Slow-freezing, Ethylene glycol, Sucrose
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