Poly-ICLC administration promotes the infiltration of Type-1 Ag-specific T cells into CNS rumor sites. (A-C), mice bearing day 10 M05 tumors were immunized with either: OVA257–264vaccines plus poly-ICLC, OVA257–264 vaccines only, mock vaccines plus poly-ICLC, or control mock vaccines on days 10 and 15. In the experiments depicted in (A-b, B-b, C), mice also received i.v. injections of 5 × 106 OT-I mice-derived naïve SPCs and LN cells on day 10 prior to the first immunization. On day 16, the mice were sacrificed, and BILs from tumor-bearing hemisphere were analyzed for the presence of OVA257–264-specific T-cells using TC-labeled anti-mouse CD3 mAb and PE-labeled OVA257–264-specific tetramer. N = 5 mice/group. BILs from the same group were pooled and compared between groups. (A), numbers represent the percentage of CD3+/OVA tetramer+ cells in lymphocyte-gated BILs. (B), total numbers of CD3+/OVA tetramer+ BILs per mouse with (b) or without (a) adoptive transfer of naïve OT-1 mouse-derived SPCs and LN cells. (C), expression of IFN-γ by isolated BILs (a), SPCs (b), ipsilateral inguinal (c), and cervical (d) LN cells. Aliquots of 1.0 × 106/ml isolated lymphocytes were cultured with 5 μg/ml OVA257–264 and 20 IU/ml rhIL-2 for 6 h (BILs) or 5 days (for SPCs and LN cells), and IFN-γ in the supernatant was measured by specific ELISA. For BILs (a), *P < 0.05 compared to all other groups. For SPCs (b), *P < 0.001 compared to all other groups. For iLNs (c) and cLNs (d), *P < 0.001 for the combination group compared to all other groups, #P < 0.01, for the vaccine alone group compared to the control or the poly-ICLC alone group. Columns, mean of three wells in 96 well plate; Error bars, SD. Representative of 2 and 4 independent experiments with similar results, for (a) and (b), respectively.