*2.7. Immunofluorescence Staining of Human NETs*

PMNs were fixed with 4% paraformaldehyde, permeabilized with 0.15% Triton X-100 in PBS, and blocked with 5% normal chicken serum (Sigma) in PBS. To stain NETs, slides were incubated with a mouse monoclonal anti-myeloperoxidase antibody (1:200; Santa Cruz sc-52707) and a secondary Alexa Fluor® 488-conjugated chicken anti-mouse IgG antibody (1:200; Molecular Probes A-21200). After staining of DNA with DAPI, neutrophil-derived NET formation was visualized by immunofluorescence microscopy performed on an Olympus model IX70 inverted microscope outfitted with an IX-FLA fluorescence observation system equipped with a FITC and DAPI filter cubes (Chroma Technology, Brattleboro, VT, USA) through Uplan FI objectives (20×, 60×). Images were captured by an Olympus XM10 digital camera using CellSens imaging software (Olympus America, Melville, NY, USA).
