CD157 cross-linking induces a change of cell shape and F-actin distribution in neutrophils. (A-B) Neutrophils were incubated with anti–CD157-biotin (or anti–HLA class I–biotin) and streptavidin-TRITC under the indicated conditions, fixed, permeabilized, and stained with phalloidin-FITC. Then, samples were observed by differential interference contrast (DIC) and fluorescence confocal microscopy. CD157 cross-linking progressively induces neutrophil polarization, whereas HLA class I cross-linking has no effects. (B) The effects following CD157 cross-linking were inhibited when the experiment was performed in the presence of 10 μg/mL anti-CD18, but not in the presence of anti-HLA class I mAb, used in the same experimental conditions. (C) Clustering of the CD11b/CD18 complex induces spatial colocalization of CD157. CD11b/CD18 clustering was induced by incubation of PMNs (30 minutes at 37°C) on microscope slides coated with FCS. Cells were fixed and stained with anti-CD11b-FITC, anti-CD157-biotin and streptavidin-TRITC. Cells were mounted onto slides and imaged at 20°C using a confocal scanning laser microscope (FV300) mounted on an IX71 inverted microscope (both from Olympus, Hamburg, Germany) with a PlanApo 60 × oil, 1.4 numerical aperture (NA) objective lens. Data are representative of 5 experiments with cells from different donors. Bar, 10 μm.