Journal: Cancer Science
Article Title: Administration route‐dependent induction of antitumor immunity by interferon‐alpha gene transfer
doi: 10.1111/j.1349-7006.2010.01578.x
Figure Lengend Snippet: Activation of antigen‐presenting cells isolated from the mice treated by interferon‐alpha (IFN‐α) gene transfer. (a) Expansion of tumor‐responsive T cells after intratumoral injection of recombinant adenovirus vector expressing mouse interferon‐α (Ad‐mIFN). ELISpot assay of interferon‐gamma (IFN‐γ)‐producing cells was performed in response to stimulation of CT26 cells. Ad‐mIFN or recombinant adenovirus vector expressing alkaline phosphatase (Ad‐AP) was injected into the CT26 tumor or into the tail vein and 20 days later splenocytes were co‐cultured with CT26 cells or lymphocytes, and stained with biotinylated antimouse IFN‐γ antibody to detect captured IFN‐γ (n = 3). Lymphocytes were isolated from the spleen of a naïve BALB/c mouse. (b) Expression of co‐stimulatory molecules on CD11c+ cells. Flow cytometry of CD40, CD80, and CD86 expressions was performed on CD11c+ cells isolated from regional lymph nodes of the mice treated by intratumoral or intravenous injection of Ad‐mIFN or Ad‐AP (n = 3–4). The frequency of CD11c+ cells (upper left) per lymphocytes, and CD40+ (upper right), CD80+ (lower left), and CD86+ (lower right) cells per CD11c+ cells are presented. (c) Number of IFN‐γ‐producing cells by the stimulation of CD11c+ cells. CD11c+ cells were isolated from the tumors of treated mice. CD4+ T cells were isolated from the spleens of CT26 tumor‐bearing mice, and co‐cultured with CT26 cells and the isolated CD11c+ cells for 3 days, and then the number of IFN‐γ‐producing CD4+ T cells was counted. AP‐CD11c+, CD11c+ cells isolated from tumors in mice treated by Ad‐AP. mIFN‐CD11c+, CD11c+ cells isolated from tumors in mice treated by Ad‐mIFN.
Article Snippet: The 1 × 10 6 of cells from the lymph nodes were incubated at 4°C for 30 min with phycoerythrin‐conjugated hamster antimouse CD11c monoclonal antibody (HL3; IgG; BD Biosciences), FITC‐conjugated rat antimouse CD40 monoclonal antibody (3/23; IgG; BD Biosciences), FITC‐conjugated hamster antimouse CD80 monoclonal antibody (16‐10A1; IgG; BD Biosciences), FITC‐conjugated rat antimouse CD86 monoclonal antibody (GL1; IgG; BD Biosciences), or isotype control antibody (IgG), and then washed twice with PBS containing 2% BSA.
Techniques: Activation Assay, Isolation, Injection, Recombinant, Plasmid Preparation, Expressing, Enzyme-linked Immunospot, Cell Culture, Staining, Flow Cytometry
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