The chemokine stromal cell‐derived factor‐1 (SDF‐1) that is released by bone marrow (BM) stromal cells and contributes to stem cell homing may also play a role in the trafficking of leukaemic cells. We analysed SDF‐1‐induced intracellular calcium fluxes in leukaemic blasts from the peripheral blood of patients with newly diagnosed acute myeloid leukaemia (AML) and lymphoblastic leukaemia (B‐lineage ALL), determined the effect of BM stromal cell‐conditioned medium on in vitro transendothelial migration (TM) and measured expression of the SDF‐1 receptor, CXCR4, by flow cytometry. AML FAB M1/2 blasts did not show calcium fluxes and TM was not stimulated. In myelomonocytic AML (M4/5), however, SDF‐1 induced significant calcium fluxes and TM was increased twofold by the conditioned medium. M3 and M4 blasts with eosinophilia (M4eo) showed intermediate activity and M6 blasts showed no functional activity. In ALL, strong calcium fluxes and increased TM (2.5‐fold) were observed. Accordingly, expression of CXCR4 was low in undifferentiated (M0) AML, myeloid (M1/2) AML and erythroid (M6) AML, but high [mean fluorescence (MF) > 50] in promyelocytic (M3) AML, myelomonocytic (M4/5) AML and B‐lineage ALL. We conclude that, in AML, SDF‐1 is preferentially active in myelomonocytic blasts as a result of differentiation‐related expression of CXCR4. Functional activity of SDF‐1 and high expression of CXCR4 in B‐lineage ALL is in accordance with the previously described activity of SDF‐1 in early B cells. SDF‐1 may contribute to leukaemic marrow infiltration, as suggested by increased CXCR4 expression and migratory response in BM‐derived blasts compared with circulating cells.