Isolation and characterization of an anti-peptide monoclonal antibody to human erythropoietin.
A site-specific monoclonal antibody to human erythropoietin has been developed. It is secreted by a hybridoma cell line derived from the fusion of murine myeloma cells with the splenocytes of a mouse that had been immunized with a 26-residue synthetic peptide antigen homologous to the amino-terminal sequence of the hormone. The antibody binds specifically to peptide, 125I-erythropoietin, and biologically active erythropoietin. The equilibrium dissociation constants of the antibody-erythropoietin and the antibody-peptide interactions are identical, Kd = 6.7 X 10(-9) M, suggesting strong conformational similarity or identity of the epitope as expressed on the peptide and the hormone. Immune complexes formed between the antibody and either human or rat erythropoietin exhibit full biologic activity. However, the antibody does not recognize the baboon, sheep, or canine hormones, indicating antigenic differences or structural variation among these erythropoietins. These results indicate that the amino-terminal region of erythropoietin is not involved in receptor binding. Furthermore, they form a basis for the study of the structure and function of the hormone using anti-peptide antibodies.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification*; D000936:Antigen-Antibody Complex; D003429:Cross Reactions; D004285:Dogs; D004921:Erythropoietin / Q000032:analysis*; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007457:Iodine Radioisotopes; D051379:Mice; D008807:Mice, Inbred BALB C; D010215:Papio; D051381:Rats; D012756:Sheep; D013045:Species Specificity
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D007457:Iodine Radioisotopes; D004921:Erythropoietin