Fig. 4
CR Abs-triggered NETosis is dependent on ACE2 and the Fc receptor. Isolated human neutrophils were treated with recombinant RBD protein (10 μg/mL) and the indicated antibodies (mAb 127, mAb 127-F(ab’)2, or cmIgG-F(ab’)2, 10 μg/mL) in the presence or absence of additional rACE2 (1 or 10 μg/mL) for 24 h. The supernatants were collected to measure the levels of (A, E) IL-8 and (B, F) MPO using ELISA kits. (C, D, G, H) The cell suspensions were spun onto a microscope slide by using a cytocentrifuge and were fixed and subjected to immunofluorescence staining with an anti-MPO antibody (green), anti-histone antibody (red) and Hoechst (blue) nuclear stain, and the area of NET formation was quantified by ImageJ. The averages of triplicate cultures ± SD are shown. The views for NET quantification were randomly selected with 9 pictures from each experiment. Statistical significance was calculated using one-way ANOVA and Tukey’s post hoc test; P values were displayed, and for values less than 0.0001, they were represented as ****. Bar: 25 μm