In Vitro Culture for Differentiation Simulation of Leishmania spp

In the 1990s my laboratory discovered that Leishmania promastigotes can combine two environmental cues, typical to lysosomes, acidic pH (~5.5) and body temperature (37 °C) into a single signal that induced differentiation. Based on this concept, we modified EARLS-based medium 199 to become an amastigote-specific medium. Shifting promastigotes to this medium followed by incubation in a CO₂ incubator induced differentiation. Axenic amastigotes reach maturation within 5 days, resembling the time it takes in vivo. This chapter provides a complete protocol we developed for L. donovani promastigote-to-amastigote differentiation. This protocol should be useful for both old-world and new-world species of Leishmania.