Department of Laboratory Medicine, Osaka University Graduate School of Medicine, Suita, Osaka 565-0871, Japan.
Cytometry A. 2010 Nov;77(11):1032-7. doi: 10.1002/cyto.a.20973.
Recent studies using stem cells or cancer stem cells have revealed the importance of detecting minor populations of cells in blood or tissue and analyzing their biological characteristics. The only possible method for carrying out such procedures is fluorescence-activated cell sorting (FACS). However, FACS has the following two limitations. First, cells without an appropriate cell surface marker cannot be sorted. Second, some laborious procedures such as rapid sorting or treatment under sterilized conditions may require in order to analyze their biological characteristics. If a specific mRNA in a particular cell type can be stained with a florescent dye and then the cells can be sorted without causing RNA degradation, a more simple and universal method for sorting and analyzing cells with a specific gene expression pattern could be established since the biological characteristics of the sorted cells could then be determined by analyzing their gene expression profile. In this study, we established a basic protocol for messenger RNA quantification after FACS (FACS-mQ) using a cRNA probe. This method could be used for the detection and analysis of stem cells or cancer stem cells in various tissues.
最近使用干细胞或癌症干细胞的研究揭示了检测血液或组织中细胞的小群体并分析其生物学特性的重要性。进行此类操作的唯一可能方法是荧光激活细胞分选 (FACS)。然而，FACS 有以下两个限制。首先，没有适当的细胞表面标志物的细胞不能被分选。其次，为了分析它们的生物学特性，可能需要一些繁琐的程序，例如快速分选或在无菌条件下处理。如果可以用荧光染料对特定细胞类型中的特定 mRNA 进行染色，然后可以在不引起 RNA 降解的情况下对细胞进行分选，则可以建立一种更简单、更通用的分选和分析具有特定基因表达模式的细胞的方法，因为可以通过分析其基因表达谱来确定分选细胞的生物学特性。在这项研究中，我们使用 cRNA 探针建立了 FACS 后信使 RNA 定量的基本方案 (FACS-mQ)。该方法可用于检测和分析各种组织中的干细胞或癌症干细胞。
