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대한생식의학회지 제31권 제1호 2010년
생쥐 난자의 효율적인 냉동보존 방법 확립을 위한 연구
삼성제일병원 생식생물학 및 불임 연구실
최수진, 김수경,김지선,조재원,전진현,변혜경
Development of Effective Cryopreservation Method for Mouse Oocytes
Su Jin Choi, Soo Kyung Kim, Ji Sun Kim, Jae Won Cho, Jin Hyun Jun, Hye Kyung Byun
Laboratory of Reproductive Biology and Infertility, Samsung Cheil Hospital and Women's Healthcare Center, Seoul, Korea
Objective: The purpose of this study was to evaluate the efficacy and effect of various cryopreservation method on the survival and the cytoskeletal stability of metaphase II mouse oocyte. Methods: Mouse ovulated oocytes were collected and cryopreserved by a modified slow-freezing method with 1.5 M 1,2-propanediol (PrOH)+0.1 M sucrose or by vitrification using cryo loop and EM grid with 40% ethylene glycol+0.6 M sucrose. Four hours after thawing, intact oocytes were fixed and stained with fluorescein isothiocyanate (FITC)-conjugated monoclonal anti-β-tubulin antibody to visualize spindle and propidium iodide (PI) to visualize chromosome. Spindle morphology was classified as follows: normal (barrel-shaped), slightly and absolute abnormal (multipolar or absent). Results: Survival rate of the frozen-thawed oocytes in vitrification group was significantly higher than that of slow-freezing group (62.7% vs. 24.4%, p<0.01). Vitrification with cryo loop showed significantly higher survival rate than that with EM grid (67.7% vs. 53.5%, p<0.05). On the other hand, proportion of normal spindle and chromosome configurations of the frozen-thawed oocytes between two vitrification group was not significantly different. Conclusion: For mouse ovulated oocytes, vitrification with cryo loop may be a preferable procedure compared to slow-freezing method. Further study should be needed to investigate developmental competency of frozen-thawed mouse oocytes.
키워드 : Mouse oocyte, Vitrification, Slow-freezing, Spindle, Chromosome
교신저자 : byun0314@unitel.co.kr
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