Simultaneous multicolor flow cytometric detection and analysis of IL-17A and IFN-γ mRNA and protein levels expressed in human T lymphocytes (P6364)

Abstract

Abstract The ongoing development of multiparameter flow cytometers, fluorescent dyes and specific antibodies for surface and intracellular markers provides valuable tools to analyze and define cellular and molecular mechanisms that underlie complex biological systems. These tools have contributed significantly to both basic and clinical research applications including immune system monitoring, drug screening, and diagnostic tests. Recently-developed techniques and reagents allow for the specific, amplified fluorescent detection of different RNA transcript levels expressed in individual cells. In this study, we evaluated the use of multicolor flow cytometry for the simultaneous analysis of differentially coexpressed mRNA and protein levels in primary cells. Using a human Th17-like cell model system, we demonstrate that in vitro-differentiated Th17-like cells can be simultaneously probed for their differentially expressed levels of IL-17A and IFN-γ proteins as well as IL-17A and IFN-γ mRNAs and IL-23 receptor mRNA. These results suggest that numerous potential new mRNA biomarkers can now be defined and used along with conventional immunofluorescent staining of protein biomarkers, to more fully characterize the nature of cells that function in complex biological systems. Simultaneous multicolor flow cytometric analysis of mRNA and protein in individual cells constitutes an important new toolset that can provide novel information from basic and clinical research investigations.