The pathogenic human parvovirus B19 has extreme tropism for human erythroid progenitor cells and has resisted cultivation in conventional cell lines. We report first propagation of this virus in an erythropoietin-dependent strain of a megakaryoblastic leukemia cell line called UT-7. Virus protein was present in about 5% of cells after 1 week of culture. Appropriate ratios of major and minor capsid proteins were determined by immunoblot, and newly synthesized capsid protein was detected by immunoprecipitation of radioactively labeled cell lysates. High molecular weight monomer and dimer intermediates were detected by Southern analysis, indicating active viral replication. Approximately 1,000 genome copies were present per infected cell, and at the optimal multiplicity of infection 20- to 50- fold more virus was produced than inoculated. Virus propagation only occurred in UT-7 cells that were adapted to growth in erythropoietin; virus signal was not detected in UT-7 cells adapted for growth in granulocyte-macrophage colony-stimulating factor or interleukin-3, even with exposure to erythropoietin for several days. Infectious virus was detected in cultures as long as 3 months after inoculation. Despite persistence, there was no evidence of viral integration on Southern analysis. This cell line may prove useful for the production of infectious virus and in the analysis of B19 parvovirus persistence, cytotoxicity, and permissivity.