Apc anti cd66b

1x10^6 PMNs were incubated with the following surface antibodies for 20 min on ice in the dark. Surface antibodies included PerCP-Cy5.5-anti-CD11b, APC-anti-CD63, APC-anti-CD66b (all Biolegend). To exclude dead cells, a fixable viability dye was included. Flow cytometry was performed using an LSR II (BD Bioscience) and analyzed with FlowJo (TreeStar).

100 µl whole blood was incubated in human Fc Receptor Block (Biolegend) followed by anti-human fluorophore-conjugated antibodies: PerCP/Cyanine5.5 anti-CD3), APC/Cyanine7 anti-CD11b, Brilliant Violet 650 anti-CD25, PE/Cyanine5 anti-CD62P/P-Selectin, Brilliant Violet 711 anti-CD45, APC anti-CD66b, Brilliant Violet 605 anti-CD56/NCAM, Alexa Fluor488 anti-CD14, Brilliant Violet 785 anti-CD8, PE anti-CD16 (all from Biolegend); Alexa Fluor594 anti-ACE-2 (R&D Systems), PerCP-eFluor710 anti-CD19 (eBioscience), and BV421 anti-HLA-DR, DP, DQ (BD Biosciences). Red blood cells were lysed and cells were fixed using 1-step Fix/Lyse Solution (eBioscience), rinsed with FACS buffer, and resuspended in FACS buffer (0.5% BSA, 0.005% EDTA, 1 × PBS). Samples were analyzed using the NovoCyte Quanteon flow cytometer, NovoSampler Q, and NovoExpress Software. An average of 1 × 10⁵ events were collected from the “Live Cell” gate for analysis. Analysis was conducted using FlowJo software version 10. The complete gating strategy is provided in Supplemental Fig. 2.