Supporting information for Dorner et al. (April 23, 2002) Proc. Natl. Acad. Sci. USA, 10.1073/pnas.092141999.
Fig. 6.
Intracellular flow cytometry detection of murine activation-induced, T cell-derived, and chemokine-related cytokine (ATAC), macrophage inflammatory protein (MIP)-1a, MIP-1b, and regulated on activation normal T cell expressed and secreted (RANTES) in activated natural killer (NK) and T cells. (A) Specificity of staining. C57BL/6 splenocytes were stimulated with phorbol 12-myristate 13-acetate, ionomycin, and brefeldin A for 6 h, stained with an anti-CD8 mAb, fixed, and stained for intracellular ATAC, MIP-1a, MIP-1b, or RANTES. The black profiles indicate the background signals obtained after blocking with an excess of unlabeled mAbs (in the case of ATAC) or after preincubation of the chemokine-specific antisera with excess amounts of their respective recombinant ligands before staining, as described in Materials and Methods. Shown are the results after gating on CD8+ cells in a representative experiment out of six. (B and C) Expression of ATAC, MIP-1a, MIP-1b, and RANTES in activated NK and CD8+ T cells and CD4+ T cells. C57BL/6 splenocytes were stimulated as described for A, stained with the indicated surface markers, fixed, counterstained with the cytokine/chemokine-specific reagents, and analyzed by two-color flow cytometry. The figures in the right upper quadrants indicate the percentages of DX5+, CD8+, or CD4+ cells positive for the given cytokine or chemokine. Shown is a representative experiment out of six.