Xenograft Tumor Growth Monitoring and Immunotherapy

All of the animal procedures used were approved by the institutional animal care and use committee at the University of Chicago. Nude mice were purchased from Harlan Sprague-Dawley. C57BL/6 mice were obtained from Envigo. NSG (severely combined immunodeficient (NOD/SCID) interleukin-2 receptor (IL-2R) gamma chain null) mice were obtained from Jackson Laboratory. For xenograft experiments, one million cells were injected subcutaneously into the right flanks of 6-week-old female nude or C57BL/6 mice. Tumor growth was monitored and measured weekly by a caliper, and tumor volume was calculated using the formula, Tumor volume (mm³) = d² × D/2, where d and D are the shortest and the longest diameters, respectively. For treatment with anti-PD-1 antibody (BioXCell, clone RMP1-14) or isotype control IgG antibody (BioXCell, clone 2A3), B16F10 melanoma cells (5 × 10⁵) were inoculated subcutaneously into C57BL/6 or NSG mice. When the tumors reached a volume of 80–100 mm³, mice were treated with anti-PD-1 or isotype control antibody (200 μg/mouse) by i.p. injection, every other day for three times. For IFNγ blockade treatment, C57BL/6 mice were treated with anti-IFNγ antibody (BioXcell, Clone XMG1.2) or isotype control IgG (BioXcell, Clone HRPN) (250 μg/mouse) every other day after tumor cell inoculation^{50 (link),51 (link)}.