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Abstract Background Recent renewed emphasis on the eradication of malaria has highlighted the need for more tools with which to achieve this ambitious goal. One high priority area is the need to determine the gametocytocidal activity of both currently used anti-malarial drugs and those in the development pipeline. However, testing the activity of compounds against Plasmodium falciparum gametocytes is technically challenging both in vivo and in vitro. Methods Here the use of a simple robust assay to screen a panel of currently used and experimental anti-malarial drugs against mature P. falciparum gametocytes is described. Results Eight of 44 compounds tested reduced gametocyte viability by at least 50% and three showed IC50 values in nM range. Conclusions There is a need to identify new compounds with activity against late stage gametocytes and the information provided by this in vitro assay is a valuable first step, which can guide future clinical studies.