期刊
CRYOBIOLOGY
卷 56, 期 3, 页码 223-232出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.cryobiol.2008.03.005
关键词
cryopreservation; vitrification; 1,2-propanediol; trehalose; murine ES cells; quartz micro-capillary; plastic straw; open-pulled straw; French-type straw; cooling rate
资金
- NIBIB NIH HHS [R01 EB002340-05, EB002340, R01 EB002340] Funding Source: Medline
Conventional cryopreservation protocols for slow-freezing or vitrification involve cell injury due to ice formation/cell dehydration or toxicity of high cryoprotectant (CPA) concentrations, respectively. In this study, we developed a novel cryopreservation technique to achieve ultra-fast cooling rates using a quartz micro-capillary (QMC). The QMC enabled vitrification of murine embryonic stem (ES) cells using an intracellular cryoprotectant concentration in the range used for slowing freezing (1-2 M). The cryoprotectants used included 2 M 1,2-propanediol (PROH, cell membrane permeable) and 0.5 M extracellular trehalose (cell membrane impermeable). More than 70% of the murine ES cells post-vitrification attached with respect to non-frozen control cells, and the proliferation rates of the two groups were similar. Preservation of undifferentiated properties of the pluripotent murine ES cells post-vitrification cryopreservation was verified using three different types of assays: the expression of transcription factor Oct-4, the presentation of the membrane surface glycoprotein SSEA-1, and the elevated expression of the intracellular enzyme alkaline phosphatase. These results indicate that vitrification at a low concentroition (2 M) of intracellular cryoprotectants is a viable and effective approach for the cryopreservation of murine embryonic stem cells. (C) 2008 Elsevier Inc. All rights reserved.
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