Leishmaniasis is a parasitic infection caused by several species of the genus Leishmania that is considered as a neglected disease. Drug development process requires a robust and updated high-throughput technology to the evaluation of candidate compounds that imply the manipulation of the pathogenic species of the parasite in the laboratory. Therefore, it is restricted to trained personal and level II biosafety environments. However, it has been established the utility of Leishmania tarentolae as a model for in vitro screening of antileishmanial agents without the necessity of level II biosafety setups. In parallel the transfection of Leishmania parasites with reporter genes as the eGFP using non-commercial integration vectors like the pIRmcs3(-) has proved to be a powerful tool for the implementation of semi automatized high-throughput platforms for the evaluation of antileishmanial compounds. Here we report the generation of a new L. tarentolae strain overexpressing the eGFP gene harboured by the non-commercial vector pIR3(-). We also demonstrate its utility for the semi-automatized screening of antileshmanial compounds in intracellular forms of the L. tarentolae parasite.