Grimm et al. BMC Cancer 2010, 10:82 http://www.biomedcentral.com/1471-2407/10/82 Page 6 of 11 Figure 2 (a) Immunohistochemistry and (b) immunofluorescence double staining: Representative image of CD4 (red)/CD8 (blue) (top) expression and Foxp3 (FITC)/CD4 (Cy3) (bottom) expression in liver metastases on day 10 (n = 7) compared to increased CD4 (red)/ CD8 (blue) expression and Foxp3 (FITC)/CD4 (Cy3) expression on day 20. Immunohistochemistry: Nova red brick red color, Vector Blue blue color. Haemalaun blue color-nuclear counterstaining. Asterisk with indication lines show metastases next to normal tissue (×100 and ×400). Immunofluorescence: FITC green fluorescein isothiocyanate, Cy3 red and DAPI 4’,6-diamidino-2-phenylindole blue - nuclear counterstaining (×250). Figure 3 Real Time PCR: Significantly increasing gene expression of IL-10, TGF-b, TNF-a but a decrease of IFN-g at metastatic sites in the liver on day 10, 15 and 20 after intraportal injection of 1 × 105 CT26.WT cells. Analysis was measured to control tissue (liver). Lines with asterisks show significant differences during metastatic growth.