Purified CD34⁺ haematopoietic progenitor cells were cultivated with stem cell factor, interleukin 3 (IL‐3), granulocyte–macrophage colony‐stimulating factor (GM‐CSF) and granulocyte CSF (G‐CSF) for 7 d, and thereafter non‐adherent cells were divided into two groups. Cells in one group (group A) were further cultivated for 7 d with four cytokines, and cells in the other group (group B) were further cultivated for 7 d with G‐CSF alone. On day 14, 220‐fold and 130‐fold increases in the numbers of non‐adherent cells were achieved for groups A and B respectively. These cell preparations contained 65% granulocytes for group A and 95% granulocytes for group B. These cells gained the ability to respond effectively with chemotaxis, phagocytosis and superoxide (O₂⁻) release. Cells in group B were appropriately primed by G‐CSF, GM‐CSF, tumour necrosis factor α and IL‐1β for enhanced release of . The responsiveness of these cells was identical to that of peripheral blood neutrophils, indicating that cells in group B may be in the resting state. In contrast, cells in group A were not primed by these cytokines for enhanced release of O₂⁻ and released a large amount of O₂⁻ spontaneously, indicating that cells in group A may be in the activated state. These findings indicate that mature neutrophils with normal functions were expanded ex vivo in group B and suggest that these cells could be used for possible autologous neutrophil transfusion to prevent bacterial infections during severe neutropenia after cytotoxic chemotherapy.