Anti il 10 jes5 16e3

For cell surface marker analysis, cells were stained with antibodies in PBS containing 0.1% (wt/vol) BSA and 0.1% NaN₃. The following mAbs were used in the present study: anti-CD11b (M1/70; eBiosciences, Ben Lomond, CA, USA), anti-Gr1 (RB6-8C5; eBiosciences), anti-CD45 (TU116; BD Biosciences, San Diego, CA, USA), anti-GR (EPR4595; Abcam, Cambridge, MA, USA), anti-HIF1α (IC1935P; R&D system, Minneapolis, MN, USA), anti-human HLADR (L243; Biolegend, San Diego, CA, USA), anti-human CD33 (P67.0; Biolegend) and anti-human CD11b (ICRF44; Biolegend).
Intracellular staining was analysed by FCM as described.^{18 (link)} Cells isolated from different organs were re-stimulated with LPS (L2630; Sigma, St Louis, MO, USA) for CD11b⁺Gr1⁺ cell analysis for 5 h, and GolgiStop (554724; BD Bioscience) was added for the last 2 h. After surface staining and washing, the cells were immediately fixed with cytofix/cytoperm solution (554714; BD Biosciences) and were stained with anti-TNFα (MP6-XT22; eBiosciences) and anti-IL-10 (JES5-16E3; eBiosciences). For transcriptional factor analysis, after surface staining, the cells were fixed with fixation/permeabilization buffer (00-5523; eBioscience) and stained with anti-GR and anti-HIF1α. FCM data were acquired on a FACSCalibur (Becton Dickinson, San Diego, CA, USA), and data were analysed with FlowJo (TreeStar, San Carlos, CA, USA).