A New Vitrification Method of Embryos in Bovine--closed pulled Straw Vitrification
To improve the freezing efficiency and avoid the contamination of liquid nitrogen in early bovine embryos, this study developed a vitrification method--the closed pulled straw (CPS), which has the advantage of isolating the cryoprotectant medium containing embryos from liquid nitrogen.A 0.25mL French straw was rolled and pulled until the inside diameter become 0.8 mm and wall thickness reached 0.07 mm.The straw was cut at the centre.The embryos were loaded individually into CPS as follows: a column of about 2 mm vitrification solution, air bubble, 1 to 2 uL vitrification solution containing embryos, air bubble, again about 2 mm column of vitrification solution and finally 2 mm air.Top end of CPS with air were sealed by a pre-hot forceps in flame and then the CPS were submerged into the liquid nitrogen directly.Bovine morulae and early blastocysts produced in vitro and in vivo were frozen by slow cryopreservation, OPS vitrificaton and CPS vitrification, thereinto the former two methods were the control.Morphology of post-thaw embryo was evaluated, and then good embryos were selected for successive culture for 72h, surviving rates was recorded at 24h and 72h respectively.For in vitro produced embryos, no differences in post-thaw morphologically normal embryo rates (87.9±5.2 vs.85.4±4.9, P>0.05), surviving rates at 24h (58.0±6.8 versus 56.3±4.4, P>0.05) and 72h (35.2±6.0 versus 34.9±6.7, P>0.05) were observed between OPS and CPS groups.However, both OPS and CPS vitrification groups resulted in higher post-thaw good embryo rates,surviving rates at 24h and 72h than slow freezing (75.8±6.1, 46.9±3.7 and 24.7±5.4, resp., P<0.05).For in vivo derived embryos, CPS vitrification yielded the almost same proportion of morphologically normal embryos after thawing with OPS vitrification and slow freezing (86.3±5.5 versus 88.6±7.1 and87.7±7.1, P>0.05), and also supported the higher post-thaw surviving rates at 24h (63.4±8.6 versus52.3±2.6, P<0.05), and 72h (37.2±3.8 versus 27.5±5.7, P<0.05) than slow freezing, respectively.There was no significant difference between CPS vitrification and OPS vitrification groups for surviving rates at 24h (P>0.05) and 72h (P>0.05).These data demonstrated that the CPS method could be used to cryopreserve early bovine embryos derived in vitro and in vivo.
bovine embryo cryopreservation OPS CPS slow freezing
Xueli Yu Yinghua Li Wen Deng Fengjun Liu Xiaoxia Li Linsen Zan
College of Animal Science, Henan university of Science and Technology, Luoyang,Henan Province 471003 College of Animal Science, Northwest Agriculture and Forestry University,Yangling, Shannxi Province
国内会议
陕西杨凌
英文
287-294
2011-05-01（万方平台首次上网日期，不代表论文的发表时间）