We studied the generation of human natural killer (NK) cells from CD34+ cells that were isolated from peripheral blood stem cells (PBSC) mobilized by granulocyte colony-stimulating factor (g-CSF). The isolated CD34+ cells were cultured in the presence of a combination of interleukin-1 (IL-1alpha), IL-2, and stem cell factor for 5 weeks without marrow stroma. We found that the CD34+ cells isolated from G-CSF-mobilized PBSC (G-CSF/PBSC) could differentiate into a population of NK cells which were CD56+(bright)/CD3- and showed morphologic characteristics of large granular lymphocytes. Immunophenotypic analysis of the NK cells thus generated showed that a small proportion of them expressed CD2, CD8 and CD16 surface markers and approximately half of them coexpressed CD7. This NK population exhibited cytotoxic activity against a NK-sensitive cell line, K562. These observations suggest that CD34+ cells from G-CSF/PBSC contain precursors of NK cells that can differentiate into functional NK cells.