Anti cd41a fitc

One hundred microliters of whole blood were stained with anti-CD3-PECy7 (BioLegend, San Diego, CA, USA; clone HIT3a), anti-CD8-PECy5 (BioLegend; clone SK1), anti-PD-L1-PE (BioLegend; clone 29E.2A3), anti-CD14-PECy7 (BD Bioscience, San Jose, CA, USA; clone M5E2), anti-CD41a-FITC (Immunotools, Friesoythe, Germany; clone HIP8), and anti-CD62P-APC (Immunotools; clone HI62P). The cells were then incubated for 15 min in the dark before adding 2 mL of BD FACS lysing solution 1X (BD Bioscience) for 10 min. Finally, the cells were washed with 2 mL of PBS 1X and resuspended in 400 µL of PBS 1X before acquisition by flow cytometry (MACSQuant Analyzer 10 flow cytometer; Miltenyi Biotec, Bergisch Galdbach, Germany). Negative gates for each marker were defined by fluorescence minus one (FMO) controls.

PBMCs (1 × 10⁶ cells) were incubated with anti-CD19-PEDy647, anti-CD38-PE, anti-CD41a-FITC, anti-CD41a-PE (Immunotools, Friesoythe, Germany), anti-CD19-PECy7, anti-CD27-APC, anti-BAFFR-FITC, anti-TACI-PE (BioLegend, San Diego, USA), anti-CD86-PE, anti-IgG-PE, anti-IgD-FITC (BD), anti-IgA-FITC, and anti-IgM-PE (Dako, CA, USA) mAbs and the corresponding isotype controls. Whole blood (100 μl) was incubated with anti-CD4-PECy7 (BioLegend), CD5-PE (BD Biosciences), CD19-PEDy647, and CD41a-FITC (Immunotools). Red blood cells were lysed, and white cells fixed using BD FACS lysing solution (BD Biosciences) to be analyzed by flow cytometry.