To assess the function of IgG subclass antibodies we studied the opsonization of Staphylococcus aureus (STAW) and Haemophilus influenzae type b (Hib) by natural IgG1 and IgG2 antibodies from five healthy individuals. Phagocytosis by polymorphonuclear leukocytes (PMN) was analyzed using FITC-labeled bacteria and flow cytometry. All PMN donors were typed for the High- and Low-Responder phenotype of the human Fc gamma receptor (hFc gamma) type IIa (CD32) and the NA1/NA2 allotype of the hFc gamma RIIIb (CD16). When PMN were used that were heterozygous for hFc gamma RIIa and hFc gamma RIIIb, phagocytosis of STAW opsonized with IgG1 antibodies was similar to that with IgG2, both in the presence and absence of a source of complement (agammaglobulinemic serum). Phagocytosis of Hib opsonized with IgG2 anti-Hib proved significantly lower (p < 0.05) than with IgG1 anti-Hib using preparations from 4 of the 5 individuals tested. IgG2 anti-Hib from one donor, however, proved more effective than IgG1 anti-Hib. The properties of PMN with hFc gamma RIIaLR,LR and hFc gamma RIIaHR,HR were compared, using hFc gamma RIIIb NA1/NA2-matched PMN. hFc gamma RIIaHR,HR PMN were virtually incapable of phagocytosing STAW and Hib opsonized with IgG2 anti-bodies without complement, in contrast to hFc gamma RIIaLR,LR PMN. Phagocytosis of IgG2-oposonized bacteria by hFc gamma RIIaLR,LR PMN was effectively inhibited by mAb against hFc gamma RIIa (IV.3). IgG1-mediated phagocytosis was blocked by mAb against hFc gamma RIIIb (CLB/FcRGran 1) to a greater extent than by anti-hFc gamma RIIa mAb. Inhibition studies with mAb against CR3 (B2.12) and hFc gamma RIIa showed that both receptors cooperated in phagocytosis. We conclude that PMN phagocytosis can be effectively mediated by antibacterial IgG2 anti-STAW and anti-Hib. Thus, IgG2 may have a critical role in the immune defense against these bacteria. However, the role of antibacterial IgG2 in opsonization and phagocytosis may be limited in individuals homozygous for hFc gamma RIIaHR.