Pe donkey anti rabbit igg poly4064

For confocal staining, 7 μm-thick sections of spleens were stained using Alexa Fluor^® 488 conjugated anti-CD4 (GK1.5) (BioLegend, San Diego, CA), phycoerythrin (PE)-conjugated anti-IL-17 (eBio17B7), allophycocyanin (APC)- conjugated STAT3 (M59-50) (eBiosciences, San Diego, CA), phycoerythrin (PE)-conjugated p-STAT3 Y705 (4/p-STAT3), and S727 (49/p-STAT3) (all from BD PharMingen, San Diego, CA), allophycocyanin (APC)- conjugated CD25 (PC61) (BioLegend), phycoerythrin (PE)-conjugated anti-Mouse/Rat forkhead box P3 (Foxp3) (FJK-16s) (eBiosciences), phycoerythrin (PE)-conjugated Mouse anti-STAT5 (pY694) (47/Stat5(pY694) (BD PharMingen), STAT5 (3H7) Rabbit mAb (Cell Signaling), and PE donkey anti-rabbit IgG (Poly4064) (BioLegend). Stained sections were examined under a microscope (LSM 510 Meta; Carl Zeiss, Oberkochen, Germany) at 400x magnification.

Lipopolysaccharide (LPS) from Escherichia coli O111:B4 (Sigma-Aldrich, St. Louis, MO, USA) was used. Phorbol 12-myristate 13-acetate (PMA), ionomycin, bovine serum albumin (BSA), anti-BSA rabbit IgG and anti-RASAL3 rabbit IgG were also purchased from Sigma-Aldrich. SYTOXRed, DAPI (4’,6-diamidino-2-phenylindole) and CellROX™Green were purchased from Thermo Fisher Scientific (Waltham, MA, USA). A Cytofix/Cytoperm kit with GolgiStop™ was purchased from BD Bioscience (Franklin Lakes, NJ, USA). Anti-CD45 (30-F11), anti-CD11b (M1/70), anti-Ly6G (1A8), anti-CD16/CD32 (2.4G2) (93) and PE donkey anti-rabbit IgG (Poly4064) were all purchased from Biolegend (San Diego, CA, USA). The isotype-matched control for each antibody was purchased from the same manufacturers.