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Polyfunctional cytokine production analysis of tetramer⁺ Mtb-specific CD8 T-cells.

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posted on 2013-02-21, 05:57 authored by Nadia Caccamo, Giuliana Guggino, Serena Meraviglia, Giuseppe Gelsomino, Paola Di Carlo, Lucina Titone, Marialuisa Bocchino, Domenico Galati, Alessandro Matarese, Jan Nouta, Michel R. Klein, Alfredo Salerno, Alessandro Sanduzzi, Francesco Dieli, Tom H. M. Ottenhoff

Peripheral blood mononuclear cells (PBMC) were stimulated with the same individual peptides as those present in tetramers and were stained with mAbs to CD8, IFN-γ and IL-2, or with isotype-control mAbs. After gating on CD8⁺ cells, the percentage of cells expressing IFN-γ and IL-2 was determined. (A) Representative intracellular cytokine staining data in one subject with LTBI, one TB patient before therapy and one PPD⁻ healthy donor. Numbers in the corners indicate the percentage of CD8⁺ cytokine-positive cells in each quadrant. (B) Summary cumulative data of the IFN-γ and IL-2 secretion capability of tetramer⁺ Mtb-specific CD8 T-cells in LTBI subjects (white bars) and TB patients with active disease before therapy (black bars). The data are expressed as the percentage of CD8⁺ T-cells that are IFN-γ⁺/IL-2⁻ or IFN-γ⁺/IL-2⁺. The values reported are the mean percentage of the different subset analysed for each group tested ± standard deviations (SD). *p<0.001 and **p<0.01 when compared to values in LTBI subjects.