Anti ctla 4 uc10 4b9

Tamoxifen was purchased from Sigma-Aldrich (St. Louis, MO). Carboxyfluorescein succinimidyl ester (CFSE) and LIVE/DEAD Fixable Dead Cell stain was purchased from Molecular Probes, Invitrogen (Carlsbad, CA). Antibodies were purchased from either BD Pharmingen (San Diego, CA): anti-CD4 (RM4-5), anti-CD25 (PC61), anti-CD3 (2C11), and anti-CD28 (37.51); Biolegend (San Diego, CA): anti-CD45.2 (104), anti-CD45.1 (A20), and anti-CD4 (GK1.5), or eBioscience (San Diego, CA): anti-Foxp3 (FJK-16s), anti-CD69 (H1.2F3), anti-glucocorticoid-induced TNFR family related gene (GITR) (DTA-1), and anti-CTLA-4 (UC10-4B9).

The following monoclonal antibodies were used for multiparameter flow cytometry: anti-CD4 (RM4-5, Biolegend), anti-CD25 (PC61.5, eBioscience), anti-CCR6 (29-2L17, Biolegend), anti-CD103 (2E7, Biolegend), anti-Ly5.1 (A20, Biolegend), anti-Ly5.2 (104, Biolegend), anti-CTLA4 (UC10-4B9, eBioscience), anti-Ki67 (solA15, eBioscience), anti-Bcl-2 (BCL10C4, Biolegend), anti-ICOS (7E.17G, eBioscience), anti-GITR (DTA-1, eBioscience), anti-Helios (22F6, Biolegend), anti-Neuropilin (FAB566A, R&D systems), anti-PD1 (J43, eBioscience), anti-CXCR5 (2G8, BD Biosciences). The Foxp3 staining kit was used for staining for CTLA-4, Ki67, Helios, Bcl-2 and Bcl6. Stained cells were analyzed on FACS Canto (Becton Dickinson) and were sorted on FACS Aria (Becton Dickinson). Data were analyzed with Flowjo software.