Malaria infection of red blood cells is associated with plasminogen activation. Surface immunofluorescence and immunoprecipitation experiments, using specific polyclonal and monoclonal antibodies raised against human urokinase, demonstrate that this activity is due to the binding of host urokinase-type plasminogen activator to the surface of erythrocytes infected by mature forms of Plasmodium falciparum malaria parasites. Depletion of urokinase from the culture medium leads to the inhibition of merozoite release and the accumulation of segmenter-infected erythrocytes; this inhibition is reversed by the addition of human single-chain or two-chain urokinase. These findings are consistent with host urokinase being involved in the process of merozoite release from the red blood cell.