Messenger RNA quantification after fluorescence activated cell sorting using intracellular antigens

• Published in: Biochemical and biophysical research communications Vol. 397; no. 3; pp. 425 - 428
• Main Authors: Yamada, Hiroya, Maruo, Rie, Watanabe, Mikio, Hidaka, Yoh, Iwatani, Yoshinori, Takano, Toru
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 02.07.2010
• Subjects: Actins - genetics; Animals; Antigens - analysis; Cancer stem cell; Cell Line, Tumor; Cell Separation; Cell sorting; Cryopreservation; FACS; Flow Cytometry; Humans; Immunohistochemistry - methods; Rats; RNA Stability; RNA, Messenger - analysis; RT-PCR; Stem cell; Cancer stem cell; Flow cytometry; Cell sorting; FACS; RT-PCR; Stem cell
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2010.05.112

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.