Development of a defined medium for heterologous expression in Leishmania tarentolae.

A defined medium is essential for studying nutritional requirements of microorganisms and for selective supplementation of necessary substances. Hemin is an essential ingredient for growth of Leishmania tarentolae, but tends to precipitate in aqueous solutions without further stabilization. These aggregates disturb the measurement of the optical density or the cell density and the following downstream processing. Therefore, we were looking for stabilizing substances and established a PEG-hemin-solution, which avoided flocculation and allowed the cultivation of L. tarentolae in a medium, which we termed SFP(II) medium. With addition of RNA from Saccharomyces cerevisiae to SFP(II) medium the SFP(III) medium was established. In this medium, the specific cell division rate was increased (0.103 h(-1)) and stable for longer periods of time. The evaluation of the SFP(III) medium was done in shaker flasks by successful expression and segregation of the SAG2 protein, one of the main surface antigens of Toxoplasma gondii. With establishment and evaluation of this defined medium, the status of the Leishmania tarentolae expression system as an alternative to commonly used cell cultures is supported.