Neutralization of Erythroid Burst-Promoting Activity In Vitro With Antimembrane Antibodies

Abstract

To investigate the relatedness of soluble and pelletable vesicular erythroid burst-promoting activity (BPA) present in lymphocyte-conditioned medium (LCM), we immunized rabbits with partially purified lymphocyte plasma membranes and tested the antisera for biological and immunologic crossreactivity with LCM and its component fractions. When preincubated with IgG purified from post-immune but not from preimmune serum, BPA expression by unseparated LCM, LCM-derived pellets, and supernatants was abolished in a dose-related fashion. As little as 0.001 mg/mL post-immune IgG reduced burst formation by 50%. Antimembrane IgG crossreacted on immunoblots with multiple components of both supernatants and pellets of LCM. Crossreactivity was also seen in LCM-derived supernatants that were subjected to ultracentrifugation. Soluble BPA was adsorbed from LCM supernatants incubated with antimembrane IgG-coated Staphylococcus aureus. Conversely, incubation of purified antimembrane IgG with intact circulating lymphocytes removed BPA-neutralizing effects from the antibody preparation. Antimembrane IgG incompletely suppressed erythroid colony-forming unit (CFU-E)-derived colony formation, an effect that could not be explained by alteration in erythropoietin sensitivity or action. There was no effect of the antibody preparation on erythroid differentiation of K562 cells or on CFU granulocyte/macrophage-derived colony growth, (CFU-G/M) by human or murine bone marrow. Taken together, our findings suggest that antibodies directed against lymphocyte plasma membranes react with both soluble and vesicular BPA, and that these physically separable erythroid growth factors may share antigenic determinants.