A fast and objective assay for cell mediated intra- and extracellular killing of Leishmania promastigotes

An in vitro radiometric assay is described for the detection of cytolytic activity against the obligate intracellular parasite Leishmania enriettii. The assay system can be equally well applied to non-cellular (humoral), cellular non-phagocytic, and cellular phagocytic effector situations. Leishmania promastigote organisms are DNA-labeled with [methyl-3H]thymidine [( 3H]dThd) with high efficiency. Spontaneous label release remains very low even in non-ideal culture conditions. We have modeled a leishmanicidal situation by co-cultivating L. enriettii with starch-elicited murine peritoneal macrophages for different time periods. At an effector-to-target ratio of 1:6 a highly significant [3H]dThd release can be observed after 4 h of co-cultivation, reaching 60% after 18 h. The principle advantages of this assay system are speed, high sensitivity and objectivity. This makes it suitable for mass screening of, e.g., immunomodulatory or parasiticidal agents and equally useful in both phagocytic and non-phagocytic situations.