Antigen site densities and ultrastructural distribution patterns of red cell Rh antigens

The ultrastructural distribution pattern and antigen site density of the major Rh antigens (C, c, E, e, and D) on red blood cell ghosts sensitized with WITH IgG Rh antibodies were determined using electron microscopy and ferritin conjugated rabbit anti-human IgG. The ferritin particle distribution pattern of all the Rh antigens studied was random and ranged from relatively isolated monodispersed clusters containing less than eight ferritin particles to large localized aggregates or masses of 20 or more ferritin particles. Evidence is presented to indicate that most of the antigen clustering was due to the conjugate during staining of the cell bound IgG on the stroma preparations. The conjugate-induced antigen clustering was influenced by both the titer of the conjugate and the staining time. Whether a similar degree of antigen mobility exists in the native membrane remains to be determined. Estimates of the number of cell bound IgG molecules (number of antigenic determinants) based on ferritin particle scoring of IgG Rh antibody sensitized red blood cells under nonsaturating conditions were in the range of 20,000 to 32,000 for each of the Rh antigens. These findings are consistent with the view that the Rh antigen complex is associated with a single membrane component.