Effective cryopreservation of human embryonic stem cells by the open pulled straw vitrification method
Effective cryopreservation of human embryonic stem cells by the open pulled straw vitrification method
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DOI：
10.1093/humrep/16.10.2187
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发表时间：
2001-10-01
影响因子：
6.1
通讯作者：
Trounson, AO
中科院分区：
文献类型：
--
作者：
Reubinoff, BE;Pera, MF;Trounson, AO
背景技术背景：人胚胎干细胞（ES细胞）来源于胚泡的内细胞团，并在培养中保留早期胚胎的多能细胞的特性。该研究的目的是确定开放式吸管（OPS）玻璃化冷冻方法，这是非常有效的冷冻保存胚胎，也可能是有效的人ES细胞。方法和结果：通过OPS方法玻璃化的所有人ES细胞团块在解冻时都可以回收，并且在平板接种后产生ES细胞集落。玻璃化菌落显着较小，并显示出增加的背景分化水平与对照组相比。然而，这些不希望的影响可以通过分别额外培养菌落1天和2天来克服。玻璃化冷冻的人ES细胞培养时间延长，并保留了多能细胞的特性，包括正常的核型，转录因子Oct-4的表达和人ES细胞特有的表面标志物。当移植到SCID小鼠体内时，玻璃化冷冻的细胞产生了含有所有三个胚胎胚层衍生物的畸胎瘤。结论：OPS法玻璃化冷冻法是一种可靠、有效的人多能胚胎干细胞冷冻保存方法。
BACKGROUND: Human embryonic stem (ES) cells originate from the inner cell mass of the blastocyst, and retain in culture the properties of pluripotent cells of the early embryo. The study aim was to determine whether the open pulled straw (OPS) vitrification method, which is highly effective for the cryopreservation of embryos, might be also efficient for human ES cells. METHODS AND RESULTS: All human ES cell clumps that were vitrified by the OPS method could be recovered upon thawing, and gave rise to ES cell colonies after plating. Vitrified colonies were significantly smaller and showed an increased level of background differentiation compared with control colonies. However, these unwanted effects could be overcome by additional cultivation of the colonies for 1 and 2 days respectively. The vitrified human ES cells were cultivated for prolonged periods and retained the properties of pluripotent cells, including a normal karyotype, expression of the transcription factor Oct-4 and surface markers that are characteristic to human ES cells. When grafted into SCID mice, the vitrified cells gave rise to teratomas containing derivatives of all three embryonic germ layers. CONCLUSIONS: Vitrification by the OPS method is reliable and effective for the cryopreservation of human pluripotent embryonic stem cells.