Binding of immunoglobulin classes to subpopulations of human red blood cells separated by density-gradient centrifugation

Human erythrocytes (RBC) from whole blood were separated according to their specific densities by centrifugation on a polyvinyl-pyrrolidine- coated colloidal silica matrix (Percoll) into four major subpopulations. By indirect immunofluorescence assay, the most dense RBC subpopulation, with specific density greater than 1.110 g/ml (3%-5% of total RBC), was positive for membrane-bound immunoglobulin; the remaining, less dense subpopulations were negative. IgG was present on 85%-95%, IgM on 28%-32%, and IgA on 15%-20% of the RBC in the most dense population. When these immunoglobulins were eluted, radiolabeled, and used in binding studies with autologous RBC fractions subjected to thermal and/or enzymatic treatment, they reacted specifically with the less dense RBC subpopulations. These results suggest that previously cryptic antigens were revealed by the activity of neuraminidase on the plasma membranes of the treated RBC.