Don't Look: Growing Clonal Versus Nonclonal Neural Stem Cell Colonies
Don't Look: Growing Clonal Versus Nonclonal Neural Stem Cell Colonies
复制标题
DOI：
10.1634/stemcells.2008-0558
复制
发表时间：
2008-01-01
期刊：
影响因子：
5.2
通讯作者：
Van der Kooy, Derek
中科院分区：
文献类型：
--
作者：
Coles-Takabe, Brenda L. K.;Brain, Ian;Van der Kooy, Derek
最近的报道已经挑战了神经球测定在定量评估神经干细胞（NSC）数量方面的克隆性。我们通过培养不同标记的神经细胞的混合物，使用视频显微镜观察单个神经细胞增殖，并封装单个NSC及其后代来测试神经球测定的克隆性。当使用以10个细胞/μ l或更少接种的原代细胞时，神经球测定产生克隆集落;然而，当使用传代的NSC时，仅当以1个细胞/μ l接种时，球体才是克隆的。最重要的是，在生长阶段移动平板（用显微镜观察培养物）大大增加了非克隆集落的发生率。为了确保克隆球的形成和研究克隆球形成频率的非自主效应，将单个神经干细胞封装在琼脂糖中并增殖为克隆自由漂浮球。我们证明，克隆的神经球可以通过避免运动诱导的聚集，通过单细胞跟踪，并通过封装的单细胞。干细胞2008;26：2938-2944
Recent reports have challenged the clonality of the neurosphere assay in assessing neural stem cell (NSC) numbers quantitatively. We tested the clonality of the neurosphere assay by culturing mixtures of differently labeled neural cells, watching single neural cells proliferate using video microscopy, and encapsulating single NSCs and their progeny. The neurosphere assay gave rise to clonal colonies when using primary cells plated at 10 cells/mu l or less; however, when using passaged NSCs, the spheres were clonal only if plated at 1 cell/mu l. Most important, moving the plates during the growth phase (to look at cultures microscopically) greatly increased the incidence of nonclonal colonies. To ensure clonal sphere formation and investigate nonautonomous effects on clonal sphere formation frequencies, single NSCs were encapsulated in agarose and proliferated as clonal free-floating spheres. We demonstrate that clonal neurospheres can be grown by avoiding movement-induced aggregation, by single-cell tracking, and by encapsulation of single cells. STEM CELLS 2008;26:2938-2944