FIGURE 2.

Vaccination with B cells pulsed with EG7.OVA cell-derived DRibbles and adoptively transfer of OT-I T cells eradicated E.G7-OVA tumors. A, Experimental scheme illustrates the immunotherapy protocol. C57BL/6 mice (n=5) with preestablished EG7 tumors (previously injected with 5×10⁵ EG7 tumor cells) were treated with DRibbles injected directly into both inguinal lymph nodes and received adoptive transfer of OT-I spleen cells (1×10⁷). Two intravenous injections of 5×10⁶ DRibbles-loaded B cells or unloaded B cells or PBS were given 3 and 6 days after the first injection of DRibbles. Untreated mice served as the controls. Tumor volume (B) and percentage of survival (C) in mice bearing subcutaneous tumors were monitored over time. Lymphocytes were collected from lymph nodes and spleens of vaccinated mice on the 10th day after the first immunization. (D) The spleen cells were stained with antibodies against OT-1 clonal T-cell receptor Vβ antibodies and CD8. The expression of Vβ5.1/5.2⁺ and CD8⁺ on T cells was analyzed by flow cytometry. The percentage of divided OT-I T cells is shown as mean±SEM derived from 3 mice per group (n=3). E and F, The lymphocytes were restimulated with DRibbles in vitro. Supernatants were harvested for detection of secretory interferon (IFN)-γ by ELISA after 72 hours (E) or the intracellular IFN-γ staining were performed to determine the frequency of antigen-specific T cells after 12 hours (F). *P<0.05, ***P<0.001. Data are representative of 3 independent experiments with similar results.