Anti nk1.1 pk136 apc efluor780

Cells were washed with FACS buffer (HBSS containing 0.2% BSA and 0.05% Azide), treated with anti-FcR (2.4G2), and then stained with the following Abs. Anti-CD4 (RM4-5)-BV510, anti-CD44 (IM7)-FITC, anti-TCRβ (H57-597)-PE/Cy7, anti-RORγt (Q31-378)-BV786, anti-T-bet (O4-46)-BV421, anti-PLZF (R17-809)-AF647, anti-CCR7 (4B12)-BV786, anti-IL-17A (TC11-18H10)-AF488, anti-IFNγ (XMG1.2)-FITC, anti-IL-4 (11B11)-BV421 were purchased from BD Biosciences. Anti-NK1.1 (PK136)-APC/eFluor780, anti-CD8α (53-6-7)-APC/eFluor780, anti-B7.1 (16-10A1)-APC, anti-B7.2 (GL1)-APC, anti-CD28 (37.51)-APC, anti-Egr2 (erongr2)-APC were purchased from eBioscience/Thermo Fisher. Anti-CD24 (M1/69)-AF594 and anti-CD69 (H1.2F3)-BV421 and anti-B220 (RA3-6B2)-AF700 were purchased from BioLegend. For detection of iNKT cells, PE-CD1d tetramer loaded with PBS57 (α-galactosylceramide analogue), obtained from the NIH Tetramer Core Facility, was used. For intracellular staining, Foxp3 staining kit (eBioscience/Thermo Fisher) was used for fixation and permeabilization according to manufacturer’s instructions and then stained with Abs for 30 min at 4°C. Data were collected with a FACS Fortessa (BD Biosciences) flow cytometer and analyzed with FlowJo (version 10.8) software (Tree Star).