The requirement for multiple antibodies binding different epitopes is not due to increased numbers of Fc domains on the RBC surface. A mixture of mHEL RBCs labeled with DiI and C57BL/6 RBCs labeled with DiO was incubated in vitro with the indicated monoclonal antibody and/or F(ab′)2 fragment combinations or PBS. After 30 minutes, the mixtures were transfused into C57BL/6 mice. Peripheral blood was obtained 3 days later, stained with polyclonal anti-HEL antisera followed by fluorescently labeled anti–mouse κ light chain, and anti-HEL staining was measured by flow cytometry. This experiment was reproduced 3 times with identical results. Representative histograms are shown.