SPF ED18 eggs were delivered with CpG DNA (n = 5), non-CpG DNA (n = 4) or PBS (n = 4) in ovo and the eggs were incubated to hatch. At day 1 post-hatch, lungs, trachea, duodenum, large intestine, spleen and bursa of Fabricius were collected in OCT, sectioned and immunofluorescent assay was performed for the quantification of IgM + B cells, CD4+ cells and CD8α+ cells. The quantitative data from the immunofluorescent assay for IgM+ B cells in spleen (a) for CD4+ cells in lungs (b), bursa of Fabricius (c) and spleen (d) and for CD8α+ cells in lungs (e) are shown. The one-way ANOVA test with Bonferroni's post test for selected comparison was performed to identify group differences and the differences were considered significant at P< 0.05. The bars represent mean ± SEM.