FACS analysis of FcγRII/III surface expression in IC-challenged mice. Protein expression of the inhibitory/activating FcγRII/III receptor pair was assessed in BAL-AM cells from indicated mice obtained 2 hours after OVA/anti-OVA challenge (ICs). Mice not receiving the OVA antigen served as Ab control (Ab). (a–c) Representative results of individual mice are shown. (a) AM cells were stained with PE anti-FcγRII/III 2.4G2 mAb. Different FcγRII/III staining patterns are specifically observed in FcγRII^–/– and FcγRIII^–/– mice after IC challenge (solid line, IC) as compared with Ab control (dashed line, Ab), demonstrating inverse regulation of surface expression of inhibitory FcγRII (reduced) and activating FcγRIII (increased) by ICs. (b) AM cells from WT or C5aR^–/– mice were stained with the anti-FcγRII mAb Ly17.2 conjugated to FITC. Note the IC-induced suppression of surface FcγRII in WT but not C5aR^–/– mice. (c) In order to achieve FcγRIII specificity, AM cells were first treated with unlabeled anti-FcγRII Ly17.2 mAb (Ly17.2 blockade) followed by PE-2.4G2 mAb staining. Substantial IC-induced upregulation of surface FcγRIII was detected in WT mice but is impaired in C5aR^–/– mice.