Protective immunity is not dependent on idiotype expression on the tumor cell surface. (A) Groups of 12 mice were vaccinated subcutaneously on days −28 and −14 with DCs loaded with irradiated, mAb-coated tumor cells, Id-KLH protein plus IL-12, or DCs alone, and challenged on day 0 with either 5000 38C13 or 5000 idiotype-negative variant V2 cells. Animals were then followed for tumor growth and killed when tumors reached 1.4 cm in diameter. (B) Anti-Id antibodies are not induced by vaccination with DCs loaded with mAb-coated tumor cells. Pooled sera from mice vaccinated with Id-KLH, DC loaded with irradiated, mAb-coated tumor cells, or naive control sera were incubated for 4 hours with wild-type 38C13 (left panels) or idiotype-negative V2 cells (right panels). Bound antibodies were detected with FITC-labeled anti-IgG (left panels) or anti-IgM (right panels) antibodies (filled histograms) using flow cytometry; lines indicate isotype control staining.