TAg-specific T cells fail to eradicate lymphoma-initiating cells. (A) Nine- to 11-week-old CD19-CreER^T2 × LoxP-Tag mice were tamoxifen treated (n = 5) or left untreated (n = 3). One group was immunized IP with 1 × 10⁷ TAg⁺ cancer cells on day −7 and subsequently tamoxifen treated (n = 5). LoxP-Tag control littermates were also tamoxifen treated (n = 4). Tamoxifen was applied on days 0-4. PCRs specific for the Cre-mediated deletion and TAg RNA expression were performed from total blood cells. PCR for β-actin served as control. Data are representative for 2 independent experiments. (B) Twelve-week-old CD19-CreER^T2 × LoxP-Tag mice were tamoxifen treated (day 0-4), and indicated organs were FACS sorted for CD19⁻, CD19⁺B220⁺CD5⁻, and CD19⁺B220^lowCD5⁺ cells on days 30 to 49. Cell numbers differed between 6000 and 100 000 cells. PCRs specific for the Cre-mediated deletion and TAg RNA expression were performed. Representative data for n = 3. (C) B cells (B) or splenocytes (sp) of CD19-CreER^T2 × LoxP-Tag mice were isolated shortly after (day 4) or long after (days 29-43) tamoxifen treatment (days 0-4, 7- to 10-week-old mice) and injected into Rag^−/− mice. Each mouse received 1 × 10⁷ B cells. One group was immunized with 1 × 10⁷ TAg⁺ tumor cells 1 week prior to tamoxifen treatment (day −7, symbols in red). One group received tamoxifen after transfer in the recipient host. Shown are combined data of 2 independent experiments. Outgrowth of TAg⁺ B cells was analyzed in blood 83 days after transfer (upper graph). Kruskal-Wallis test and Dunn’s post hoc test were performed; adjusted P values are indicated. Mice with >1% TAg⁺ CD19⁺ cells in blood or spleen were considered lymphoma bearing, and lymphoma onset is depicted as Kaplan-Meier graph. Censored mice were euthanized because of other signs of distress.