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. 2013 Jun;48(6):758–764. doi: 10.1165/rcmb.2012-0499OC

Figure 1.

Figure 1.

CCR7 expression by human eosinophils. Flow cytometry events representing intact cells were analyzed, and debris was excluded by forward- and side-scatter gating. Shaded histograms represent CCR7 staining and are overlaid on unshaded dashed histograms representing irrelevant isotype control staining. (A) Flow cytometry of unstimulated, freshly purified eosinophils and IL-5–primed eosinophils for surface expression of CCR7. Eosinophils demonstrating viability by propidium iodide exclusion were included in the analysis. The histogram plots show single experiments representative of four independent experiments for unstimulated eosinophils and IL-5–stimulated eosinophils. The set of histograms displayed is from the same donor. Data are expressed as percentage of maximum cell count (% of Max). Forward scatter versus CCR7 fluorescence plots of these data are presented in Figure E2. Bar graphs depict geometric mean fluorescence intensity (MFI) averaged over the four donors tested for the freshly purified, unstimulated condition (P = 0.08, paired t test; bars represent SEM) and the IL-5–primed condition (*P < 0.03, paired t test; bars represent SEM). (B) Flow cytometry of unstimulated, freshly purified eosinophils and IL-5–primed eosinophils for total CCR7 expression. Eosinophils were fixed in 2% paraformaldehyde and permeabilized with 90% methanol before staining. The histogram plots are from a single donor and are representative of four separate experiments from four separate donors. Forward scatter versus CCR7 fluorescence plots of these data are presented in Figure E3. Bar graphs depict geometric MFI averaged over the four donors tested (*P < 0.005, paired t test; bars represent SEM). (C) Western blot for CCR7 of freshly purified, unstimulated eosinophils and IL-5–primed eosinophils. Peripheral blood mononuclear cells (PBMCs) from the donor providing eosinophils served as a positive control. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) expression served as a loading control. Data are representative of four independent experiments.