Clodronate (dichloromethylene bisphosphonate) inhibits the activity of osteoclasts, thereby preventing bone resorption in disorders characterized by excessive bone loss. Intravenously injected clodronate encapsulated in liposomes is also known to inactivate phagocytic cells in spleen and liver in vivo. The macrophage suppressive effect of clodronate is of interest in autoimmune diseases, like rheumatoid arthritis, in which phagocytic cells are involved in inflammatory processes, but knowledge of the interaction of clodronate with phagocytic cells is scarce. We have studied the uptake of clodronate, both free and encapsulated in negatively charged liposomes, by the macrophage-like cell line RAW 264 and by other types of cell lines. The uptake was assessed by a growth inhibition assay. The liposome-encapsulated clodronate was 50 and 350 times more potent than free drug for RAW 264 and CV1-P, respectively. Cell lines with a lower endocytotic capacity were insensitive to liposome-mediated delivery of the drug. The action of free clodronate seemed to be extracellular in all cell lines studied. Calcium and/or iron have been suggested to be involved in the intracellular uptake and action of clodronate in phagocytic cells. We found that the uptake of free clodronate by RAW 264 cells was indeed mediated by calcium and iron, while the uptake of liposomal drugs was only slightly affected by calcium. The increased intracellular calcium concentration in macrophages did not significantly affect the growth-inhibitory properties of clodronate, whereas iron loading of the cells partially restored the cell growth. The data do not support the role of calcium chelation as a mechanism of action of clodronate, but suggest that intracellular iron is, at least partially involved.