High efficiency plating method for Leishmania promastigotes in semidefined or completely-defined medium.

A simple technique, developed for the isolation of clones derived from single, promastigote cells of Leishmania donovani and Leishmania tropica, involved the use of semisolid agar. Both species of Leishmania promastigotes formed discrete colonies at high efficiency either in semidefined medium containing 10% fetal calf serum or in completely-defined medium lacking serum. Visible colonies appeared between 8 and 14 days in growth medium containing 10% fetal calf serum. Replacement of the fetal calf serum with bovine serum albumin and Tween-80 increased the time of colony formation by 50% but did not affect the cloning efficiency. Viability of colonies transferred from semisolid agar to liquid suspension culture was 100%.