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題 名 | 從急速冷凍卵巢再回收的豬卵之存活率=Survival Rate of Porcine Oocytes Recovered from Ultrarapid Frozen-Thawed Ovaries |
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作 者 | 吳明哲; 李秀美; 駱亞欣; 鄭翠雲; 周佳樂; | 書刊名 | 畜產研究 |
卷 期 | 32:1 1999.03[民88.03] |
頁 次 | 頁1-15 |
分類號 | 437.27 |
關鍵詞 | 豬; 卵巢; 卵; 急速冷凍; 解凍法; 體外成熟; Pig; Ovary; Oocyte; Vitrification; Thawing method; In vitro maturation; |
語 文 | 中文(Chinese) |
中文摘要 | 豬卵巢的冷凍保存研究,為癌症病患接受藥物和放射線療程前或保存健康卵巢再 植回體,提供一項實驗動物模式。試驗一:把發身前女豬卵巢的濾泡卵依其回收過程所在的 液相:濾泡液、20%胎牛血清的磷酸緩衝液,或以40% Ethylene Glycol和18% Ficoll 70 兩 種溶液混合而成的EGF4018冷凍保護液中冷凍,解凍卵透明帶完整率分別為95.7%(67/70)、 89.7%(130/145)和98.4%(186/189),以及卵丘細胞層包被在透明帶外仍完整的比率為100.0% (67/67)、73.1%(95/130)和100.0%(186/186)。試驗二:把離體四至六小時的卵巢再保 存於39℃�皕酈鷎i箱、-20℃冰箱、或-196℃液態氮桶內,經驗24小時後再回收卵巢濾泡內 的卵,其卵透明帶完整率均為100%,而卵丘細胞層包被完整率分別為70.3%(52/74)、63.3% (87/137)和20.4%(54/265)。卵在濾泡內被保存,不會因卵巢所保存的溫度而導致卵透明 帶破損,但是卵透明帶外的卵丘細胞層會因冷凍而有片塊剝離情形。試驗三:卵巢先浸泡於 EGF4018或3M DMSO 冷凍保護液10分鐘後再冷凍,當在常溫解凍時,卵丘細胞層包被在 透明帶外仍完整的比率分別為28.8%和38.6%;而在70℃ 水浴槽90秒解凍時,卵丘細胞層 包被在透明帶外仍完整的比率分別為48.7%和37.9%。試驗四:把30個卵巢直接丟入液態氮 並保存35天後才解凍,有21%(109/522)的卵仍保有完好的卵丘細胞層包被在透明帶外, 經體外成熟培養48小時後,僅剩3%(16/522)的卵是存活的,且可體外受精。試驗五:從 EGF4018冷凍保護劑浸泡後冷凍的20個卵巢回收的卵經體外成熟培養48個小時後,有 10.9%(24/221)的卵存活,且經體外受精培養後有2個發育成四細胞胚。發身前女豬的卵巢可 經急速冷凍保存一段時期後,再解凍回收其濾泡卵,卵的存活率雖低但可以體外成熟培養與 受精。 |
英文摘要 | Ovary preservation in vitro at low temperatures is a promising method for protecting cancer patients from the sterilizing effects of chemotherapy and/or radiotherapy. Pig is an animal model for this purpose on restoration of fertility to oophorectomized women patients. Exp. I: Oocytes from the ovaries of prepubertal gilts after slaughter were collected in follicular fluid, in 20% fetal calf serum in phosphate-buffered saline, or in EGF4018 solution consisting of 40% ethylene glycol and 18% Ficoll 70 as cryoprotectants for cryopreservation. After thawing, oocytes with intact zona pellucida were 95.7% (67/70), 89.7%(130/145) and 98.4%(186/189) in the above three freezing media, respectively. Also, zona-intact oocytes with enclosed cumulus cell layers were 100.0%(67/67), 73.1%(95/130) and 100.0%(186/186), respectively. Exp. II: Ovaries were collected and maintained at 25 ℃ for 4~6 hours and then preserved at 39℃ incubator, -20℃ refrigerator, or -196 ℃ liquid nitrogen for 24 hours. Follicular oocytes after storage had intact zona regardless of the storage condition of ovaries. But zona-intact oocytes with enclosed cumulus cell layers were 70.3%(52/74), 63.3%(87/137) and 20.4%(54/265), respectively, Results indicated that the preservation of ovaries at various temperatures would not cause a severe collapse of zona but cryopreservation would result in such as a condition that cumulus cell layers around the zona came off in slices. Exp. III: Ovaries were immersed in EGF4018 or 3 M DMSO cryoprotectant solution for 10 minutes prior to freezing. When EGF4018 or DMSO cryoprotected ovaries were thawed at room temperature, 28.8% and 38.6% of zona-intact oocytes had an enclosed cumulus cell layers, respectively. When EGF4018 or DMSO cryoprotected ovaries were thawed at 70 ℃ for 90 seconds, 48.7% and 37.9% of zona-intact oocytes had an enclosed cumulus cell layers, respectively. Exp. IV: A total of 30 ovaries was put directly in liquid nitrogen and preserved for 35 days, After recovery, 21%(109/522)of zona-intact oocytes had enclosed cumulus cell layers around the zona. Surviving oocytes were cultured in vitro to mature for 48 hours and only 3% (16/522) of oocytes survived to the time of in vitor fertilization, Exp. V: Twenty ovaries immersed in EGF4018 prior to vitrification were thawed for oocyte collection and 10.9%(24/221) of zona-intact oocytes surevived after 48 hours of in vitro maturation. Subsequently, two 4-cell embryos were obtained after fertilization and culture in vitro, Experiments in this study demonstrated that porcine ovaries at the prepubertal stage could be ultrarapidly frozen for a period of time and then follicular oocytes recovered for in vitro maturation and fertilization although there was a low survival rate. |
本系統中英文摘要資訊取自各篇刊載內容。