Fig. 6.

Vaccination against endoglin induces specific T-cell responses. a FACS analyses of endoglin expression on endoglin-transfected D2F2 tumor cells (mEndo⁺–D2F2). Cells were stained with PE conjugated anti-endoglin Ab. Rat IgG Ab was used as an isotype control (black line). b ELISPOT analyses of IFNγ producing cells using different stimulator cells. Splenocytes, enriched for CD8⁺ cells, were isolated from vaccinated mice and incubated for 24 h with either irradiated wild-type D2F2 cells, mEndo⁺-D2F2, or HEVc endothelial cells. The mean spot number of each group is shown (n=3, mean±SD). c T cell-specific cytotoxicity against endoglin-positive HEVc endothelial target cells. Splenocytes were isolated from vaccinated mice 10 days after tumor cell challenge. A [³⁵S]-release assay was performed at different effector-to-target cell ratios with splenocytes being re-stimulated with irradiated mEndo⁺–D2F2 cells for 5 days and [³⁵S] methionine labeled HEVc used as target cells. The data depict average ± SD of triplicate wells. Similar results were obtained in three independent experiments. d Sensitivity of mEndo⁺–D2F2 and wild-type D2F2 cells (mEndo⁻–D2F2) to CTL killing. [³⁵S] methionine labeled wild-type D2F2 or mEndo⁺ D2F2 target cells were co-incubated with effectors at E/T =1:12.5. Similar results were obtained in three independent experiments. *P<0.05 compared with control wild-type D2F2 target cells