Human monocyte/macrophage systems are extremely heterogeneous. Although many attempts have been made to define monocyte subpopulations, few antigens distinguish them. We previously reported that GPI-80, a novel glycosylphosphatidyl-inositol (GPI)-anchored protein that is expressed mainly on human neutrophils regulates neutrophil adherence and migration, and that GPI-80 is expressed on monocytes. In this study, we examined the precise distribution of GPI-80-positive monocytes using flow cytometry. Using anti-CD14 and anti-CD16 mAbs, almost all GPI-80-bearing monocytes belong to the strongly CD14-positive monocyte subpopulation. Furthermore, flow cytometric analysis of GPI-80 and other monocyte markers revealed that GPI-80 expression was high in CD11b-, CD32-, and CD64-positive monocytes. In contrast, GPI-80 expression was low in HLA-DQ-positive monocytes. These results suggest that almost all GPI-80 positive monocytes belong to a monocyte subpopulation that is superior in phagocytosis and reactive oxygen production, but inferior in antigen presentation. GPI-80 may be a useful antigen for classifying monocytes into subpopulations.