Invasion Assay of E. histolytica

This was performed as described previously (Emmanuel et al., 2015 (link)). Briefly, 75,000 E. histolytica trophozoites were pre-incubated for 3 h with 0.5% DMSO, 14 and 25 μM of tipifarnib and after 3 h, cells were re-suspended in serum-free TYI medium and loaded in the upper chamber of a Corning BioCoat Matrigel Invasion Chamber (Corning). The lower chamber contained TYI medium supplemented with 10% adult bovine serum (Sigma-Aldrich). The Matrigel Invasion Chamber was incubated at 37°C for 48 h in a GasPak EZ gas-generating anaerobe pouch system. At the end of incubation, images of trophozoites in the lower chamber were captured using a 10 × objective lens fitted in a Zeiss Axiovert A1 inverted microscope. Trophozoites that had migrated into the lower chamber were also counted using a hemocytometer. The data were obtained from three independent experiments each performed in triplicate and the percentage invasion of trophozoites was calculated and plotted by using GraphPad Prism.