Immune complexes bind preferentially to specific subpopulations of human erythrocytes

Abstract

Primate erythrocytes have complement receptors (CR1) that, both in vivo and in vitro, bind immune complexes (IC) opsonized with C3b. The present study was undertaken to determine whether the ability of human erythrocytes to bind IC is a characteristic shared by all erythrocytes. Binding of IC to erythrocytes probably involves the interaction of several C3b molecules with several CR1 clustered in small areas of the erythrocyte surface. To identify IC binding CR1 clusters, we first assessed the binding to erythrocytes of fluorescein-labeled polystyrene beads coated with monoclonal anti-CR1 antibodies (anti-CR1-beads) and second, performed IC. The binding of these ligands to erythrocytes was evaluated by immunofluorescence microscopy and flow cytometry. We found that only a fraction of erythrocytes from normal individuals bound anti-CR1-beads specifically and the percentage of erythrocytes able to bind beads increased with increasing numbers of CR1 per erythrocyte. However, the number of anti-CR1-beads bound per erythrocyte varied among cells from the same individual. We demonstrated further that the erythrocyte binding sites for anti-CR1-beads are also binding sites for opsonized IC. This was shown by demonstrating that anti-CR1-beads inhibited the binding of opsonized IC to erythrocytes and opsonized IC inhibited the binding of anti-CR1-beads to erythrocytes. Incubation of erythrocytes with opsonized IC, followed by FITC-labeled secondary antibodies, confirmed that indeed only a fraction of erythrocytes is capable of binding opsonized IC and that the binding sites for IC occupy small regions on the erythrocyte membrane. By contrast, we demonstrated that greater than 90% of erythrocytes express CR1. In conclusion, only some erythrocytes have the capacity to bind IC. Differences in the ability of erythrocytes to bind IC are probably related to differences in the clustering of CR1 in the erythrocyte membrane. Anti-CR1-beads identify erythrocyte binding sites for IC. These beads should prove useful to assess the changes that occur in the erythrocyte CR1 after exposure to IC in vivo.