The combined vaccination therapy elicited memory and therapeutic antitumor immunity. (A) For primary challenge, 10 BALB/c mice per group were vaccinated with 50 μg MCP3-sFv plasmid DNA together with cardiotoxin as in Figure 1. Control mice were injected with plasmid DNA without cardiotoxin pretreatment, with PBS, or with cardiotoxin alone, or with plasmid DNA encoding an irrelevant, HIV gp120 antigen, fused with MCP3. After 3 vaccinations, all mice were challenged with a lethal dose of 2 × 10⁵ A20 lymphoma cells by intraperitoneal injection and followed for survival for 80 days. (B) Tumor-free mice collected from the primary challenge experiments were rechallenged with 2 × 10⁵ A20 tumor cells intraperitoneally and followed for survival. Data represent combined results from 3 independent experiments. Survival differences between groups were analyzed using the log-rank test. (C) In therapeutic studies, 10 BALB/c mice per group were first inoculated with 2 × 10⁵ A20 tumor cells intraperitoneally on day 0. On days 1, 4, 8, and 18 the mice were then vaccinated with plasmid DNA encoding MCP3-fused idiotype sFv with or without cardiotoxin pretreatment 5 days before the first and last vaccination. Control mice were injected with cardiotoxin or PBS alone. Data represent combined results from 2 independent experiments.