Anti tgf β 1 2 3 1d11

CD4⁺CD25^- or CD4⁺ T cells were isolated after mechanical disruption of murine spleens by using the CD4⁺CD25⁺ regulatory T cell isolation kit (Miltenyi Biotec) or CD4⁺ BD IMag beads, respectively, via manufacturer’s instructions. B cells were isolated from murine spleens after mechanical disruption by enriching for CD19⁺ B cells labeling with CD19⁺ magnetic beads and processing with MACs columns (Miltenyi Biotec). To isolate DCs, spleens were treated with collagenase and processed using a spleen dissociation kit (Miltenyi Biotec) and subsequently labeled with CD11c⁺ magnetic beads (Miltenyi Biotec). DCs or B cells were pulsed with 1 or 2 μg/ml of CBirTox for 1–2 hours, respectively, at 37°C. After washing with PBS, DCs and B cells were cultured with CD4⁺CD25^- or CD4⁺ CBir1 Tg T cells at a ratio of 1:10 or 2:1, respectively with or without the addition of anti-TGF-β (1,2,3) (1D11) (R&D Systems), LE135, retinoic acid (Tocris), or TGF-β R1 kinase inhibitor III (Calbiochem) in complete 1640 RPMI with L-gluatmaine media, supplemented with penicillin (1U/ml), streptomycin (100 μg/ml), 2.5 μM 2-ME and 10% fetal bovine serum (FBS) (Atlanta Biologicals).