Assembly of immune complexes on the surface of ECs results in FcγRIIIb-mediated PMN recruitment. (A) Rabbit anti–mouse Ig (RbAMIg; 1:250) was perfused at 0.83 mL/minute for 5 minutes after perfusion of mAb RMAC8, after which deposition on the surface of ECs was demonstrated by staining with FITC-conjugated goat anti–rabbit-Ig by flow cytometry. The profiles are for HMEC-1 perfused with RMAC8 (filled line), RbAMIg alone (gray line), or RMAC8 followed by RbAMIg (black line). (B) PMNs were perfused at 1.5 dynes/cm² over unstimulated HMEC-1 cells precoated with mAb RMAC8 alone or with mAb RMAC8 followed by RbAMIg, or over plastic coated with mAb RMAC8 (50 μg/mL) followed by RbAMIg (50 μg/mL). Immune complexes enhanced PMN arrest when coated on plastic but not when presented by ECs (*P < .001. (C) RMAC8 + RbAMIg stimulated PMN arrest at 1.5 dynes/cm² on resting ECs infected with adE-selectin (100 MOI) and adICAM-1 (50 MOI) (*P < .005), and (D) this was inhibited by fab′ anti-FcγRIIIb (3G8) but not by anti-FcγRIIa (IV.3) on PMNs (*P < .01). (E) In contrast, RMAC8 + RbAMIg stimulated PMN adhesion at 1.5 dynes/cm² to TNFα-stimulated ECs was inhibited by anti-FcγRIIa, but not significantly by anti-FcγRIIIb (*P < .004). All values are mean + SEM of 3 experiments.