Formycin B is the most active antileishmanial agent in vitro because it is metabolized by the parasites to formycin A phosphates. The in vivo use of formycin B may be limited by its toxicity to humans due to the slight similar metabolism of the drug in human cells. The obligatory intramacrophage localization of Leishmania in man suggests that encapsulation of drugs within macrophage-directed carriers, such as human red blood cells (RBCs), might enhance the therapeutic-toxic ratio. Since uncharged formycin B would be poorly metabolized by the RBC and would diffuse from the carrier, RBCs were incubated with formycin A so that approximately 90% of the formycin A was taken up by the cells, phosphorylated by RBC enzymes to charged formycin A triphosphate, and retained in the cells. In vitro, 81% of Leishmania-infected macrophages phagocytized IgG-coated RBCs containing this active form of formycin B, and multiplication of organisms within macrophages could be suppressed by approximately 80%. The 50% effective dose of the formycin A-RBC formulation was 0.02 microM, whereas the 50% effective dose of unencapsulated drug was 0.84 microM. This report of in vitro activity of human RBCs containing formycin A and coated with IgG indicates that this formulation should be tested for antileishmanial activity in vivo.