Comparison of relevant biological assays for the determination of biological active erythropoietin

Human recombinant erythropoietin (rh EPO), a glycoprotein hormone which is an obligatory growth factor for the proliferation and differentiation of committed erythroid progenitor cells, has been purified to homogeneity and compared to human urinary erythropoietin (EPO). Erythropoietin levels will be determined and standardized by in vivo bioassays in which endogenous erythropoietin production has been reduced by hypertransfusion or hyperbaric atmosphere. The effect of EPO on the rate of red cell production or Fe59 incorporation is used as measure of its erythropoietin content. Both in vivo assay systems will be compared with respect to standardization for routine laboratory use. For in vitro characterization of EPO the classical assay for Fe59 incorporation in in vitro rat bone marrow cells will be compared to the proliferation and H3 Thd uptake by treatment with phenylhydrazin enriched erythroid progenitors from mouse spleens. (The direct physiological effect of EPO on stem cells will be shown by differences in BFUe progenitor levels). Rh EPO is jointly developed by Integrated Genetics (Boston) and Behringwerke AG (Marburg).