Figure 1

Despite unimodal distribution, CD4 levels on naïve CD4 T cells are correlated with responsiveness. A. Gating strategy used to sort NCD4lo and NCD4hi cells from six- to eight-week-old mice. Right panel shows sort profile for NCD4hi and NCD4lo cells. B. CD69 expression on anti-CD3 + anti-CD28 (3 μg/ml each) stimulated NCD4lo and NCD4hi cells 16 hours post-activation. Numbers indicate representative proportions of CD69+ cells in NCD4hi (black) and NCD4lo (grey) cells. C. Proportions of CD69+ cells in a dose–response curve with 3 μg/ml of anti-CD28 and titrating anti-CD3 concentrations as shown. (Mean ± SE, n = 4; n.s.: not significant). D. Amount of IL-2 detected 48 hours post-stimulation with anti-CD3 + anti-CD28 in culture supernatants. (Mean ± SE, n = 3; Background values shown as a line). E. 3H-Thymidine incorporation assay to measure proliferation 60 hours post stimulation with anti-CD3 + anti-CD28. (Mean ± SE of triplicate cultures, 1 of >7 experiments). F. 3H-Thymidine incorporation assay on sorted NCD4hi and NCD4lo cells cultured in vitro with IL-2 for 24 hours prior to stimulation with anti-CD3 + anti-CD28 for 60 hours. (Mean ± SE of triplicate cultures, one of three experiments). G. Proportion of trypan blue-positive cells 16 hours post-activation (Mean ± SE, n = 6, data representative of four experiments). H. Cytokine levels in supernatants of in vitro primed and recalled NCD4hi and NCD4lo cells. I. IFNγ/IL-4 ratios and mean ± SE calculated based on absorbance values. SE, standard error.