Specific precipitating antibody to human urinary erythropoietin

In the present experiments, human urinary erythropoietin was purified from the urine of a patient suffering from pure red cell aplasia, using ethanol precipitation, gel filtration on the Sephadex G-100 column and electrofocusing. The anti-human urinary erythropoietin was prepared by injecting the isolated erythropoietin with Freund's complete adjuvant into rabbits intracutaneously. The specificity of the anti-human urinary erythropoietin was ascertained by a double diffusion technique and immunoelectrophoresis, observing a single precipitation line at the alpha2-globulin region against human serum or human urinary erythropoietin (Standard B). This precipitating anti-human urinary erythropoietin inhibited completely the effects of erythropoietin (Standard B) on the heme synthesis of bone marrow cells in vitro. The precipitating antigen with the specific anti-human urinary erythropoietin, that is, erythropoietic stimulating factor of the fraction 2, had the ability to enhance heme synthesis in bone marrow cells in vitro. The specific anti-human urinary erythropoietin makes it possible to determine the serum erythropoietin levels in various diseases by means of quantitative immunoelectrophoresis.