Yamada Hiroya, Maruo Rie, Watanabe Mikio, Hidaka Yoh, Iwatani Yoshinori, Takano Toru
Department of Laboratory Medicine, Osaka University Graduate School of Medicine, Suita, Osaka 565-0871, Japan.
Biochem Biophys Res Commun. 2010 Jul 2;397(3):425-8. doi: 10.1016/j.bbrc.2010.05.112. Epub 2010 May 27.
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 limitations. First, cells without an appropriate cell surface marker cannot be sorted. Second, the cells have to be kept alive during the sorting process in order to analyze their biological characteristics. If an intracellular antigen that was specific to a particular cell type could 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) targeting intracellular antigens. This method can 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少量细胞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行方法是荧光激活细胞分选（FACS）。然而，FACS 有以下局限性。首先，没有合适细胞表面标志物的细胞无法分选。其次，为了分析其生物学特性，细胞在分选过程中必须保持存活。如果能用荧光染料对特定细胞类型特有的细胞内抗原进行染色，然后在不导致 RNA 降解的情况下对细胞进行分选，那么就可以建立一种更简单、通用的方法来分选和分析具有特定基因表达模式的细胞，因为分选后细胞的生物学特性可以通过分析其基因表达谱来确定。在本研究中，我们建立了一种针对细胞内抗原的 FACS 后信使 RNA 定量（FACS-mQ）的基本方案。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。
