Sanquin Research Academic Medical Center Landsteiner Laboratory

Amsterdam, Netherlands

Sanquin Research Academic Medical Center Landsteiner Laboratory

Amsterdam, Netherlands
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PubMed | Sanquin Research Academic Medical Center Landsteiner Laboratory
Type: Journal Article | Journal: Journal of immunology (Baltimore, Md. : 1950) | Year: 2016

Effective T cell responses entail the coproduction of IFN-, TNF-, and IL-2. Cytokine production is determined by transcriptional and posttranscriptional events. However, increased transcript levels do not always translate into protein production, and therefore simultaneous transcripts and protein measurement are essential for the appropriate analysis of T cell responses. In this study, we optimized flow cytometry-based fluorescence in situ hybridization (Flow-FISH) for IFN- to multicolor flow cytometry that allows for single-cell measurement of mRNA and protein levels. This high-throughput analysis detected Ag-specific human T cells of low frequency. We also employed Flow-FISH for single-tube analysis of IFN- transcript and protein profile to simultaneously study the responsiveness of different T cell subsets, that is, naive, effector, and memory T cells. Importantly, the simultaneous transcript and protein analysis of IFN- and of TNF- and IL-2 revealed that T cell responses consist of two types: one subtype loses mRNA expression during activation, whereas the other maintains high transcript levels throughout stimulation. High cytokine transcript levels correlated with increased protein production. Intriguingly, this mRNA

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