(A) Immunofluorescence staining of human mucosal cryosections for IRF4. Cryosections of gut specimens from control patients and IBD patients (n = 10 per group) were stained with antibodies against IRF4 (red). Nuclei were counterstained with DAPI (blue). Original magnification, ×300. (B) Quantitative assessment of IRF4-positive cells. IRF4-positive cells were counted in 7 HPFs per patient. There was a significantly increased number of IRF4-expressing cells in both CD and UC patients as compared with control patients. **P < 0.01. (C) Quantitative assessment of IRF4 and IL-6 mRNA in the lamina propria. IRF4 and IL-6 mRNA levels in mucosal biopsies were determined by quantitative PCR. A total of 16 IBD patients were analyzed. Values for IRF4 and IL-6 were strongly correlated (r = 0.81) in IBD. (D) Immunofluorescence double staining for IRF4 and CD3. Double-staining analysis was performed (green, IRF4-positive cells; red, CD3-positive cells). Original magnification, ×400 (upper panels); ×1000 (lower panels). Cells coexpressing IRF4 and CD3 appeared yellow (arrow). Staining analysis revealed that a large number of IRF4-positive cells coexpress CD3 on their surface. (E) Immunofluorescence double staining for IRF4 and CD4, CD8, and CD11c. Cells were stained using anti-IRF4 antibodies, and additional surface staining was performed using antibodies against either CD11c, CD8, or CD4. The number of positive cells was analyzed in 7 HPFs per patient (n = 6). CD11c, CD8, and CD4 single-positive cells and the number of double-positive cells were counted. Data represent mean values ± SD.