Fig. 4

T Cytokine secretion is not affected by exposure to autoclaved or live soil in an anti-CD3/anti-CD28 activation assay. On day 21, single-cell suspension from splenocytes was prepared, cells were rested overnight and then either left unstimulated or stimulated with anti-CD3/anti-CD28 antibody cocktail. CD69 expression was assessed at 24 h post stimulation using flow cytometry for (A) CD4⁺ T cells and (B) CD8⁺ T cells. (C) Supernatants from unstimulated or stimulated splenocytes were collected at 48 and 72 h and analyzed using U-Plex from Meso Scale Discovery. Each symbol represents an individual mouse, lines indicate the mean 95% C.I. (n = 4 for Unstimulated and 48 h treatment, and n = 8 for 72 h, from two independent exposure experiments). The data was analyzed with One-way ANOVA