Cd16 percp

Blood granulocytes and single-cell suspensions obtained from digested biopsies of heathy donors and IBD patients were stained with the following antibodies CD66b PE-Cy7 (#305116 clone G10F5, 1/20 dilution, BioLegend), CD16 PerCP (#302028 clone 3G8, 1/20 dilution, BioLegend), CD62L APC-Cy7 (#304814 clone DREG-56, 1/20 dilution, BioLegend), CD69 BV421 (#310930 clone FN50, 1/20 dilution, BioLegend), CD193 FITC (#310720 clone 5E8, 1/20 dilution, BioLegend), CD63 FITC (#353005 clone H5C6, 1/20 dilution, BioLegend) and CXCR4 PE (# FAB170P clone 12G5, 1/10 dilution, R&D systems) and Zombie Aqua Fixable Viability Kit 1/1000 (#423101, BioLegend) for death cells. Cells were fixed using BD Stabilizing Fixative [BD], acquired using a BD FACSCanto II flow cytometer (BD) and analyzed with FlowJO software (version 10.6.1, BD).

For performing flow cytometry, we used fluorescent-labeled monoclonal antibodies to particular surface molecules: CD45-APC/Cy7, CD3-PerCP, CD3-PE/Cy7, CD4-APC, CD4-APC/Cy7, CD8-PE/Cy7, CD25-APC, CD127-PerCP/Cy 5.5, CD14-PE/Cy7, CD19-APC, CD16-PerCP, CD56-PE/Cy7 (all Biolegend, San Diego, CA, USA). CD25-PE and HLA-DR-APC/Cy7 antibodies (Biolegend, San Diego, CA, USA) were used for evaluating of the expression of activation markers. Flow cytometry analysis was performed on FACS Canto II (BD, Piscataway, NJ, USA) machine by using FACS Diva (BD, Piscataway, NJ, USA) software. We used non-stained controls, FMO controls, and isotypic controls.