In this work, the therapeutic effect of E-64, a broad spectrum cystine protease inhibitor against Giardia lamblia excystation was studied in vitro and in vivo. Purification of cysts from heavily infected human faecal samples followed by excystation and axenic cultivation of the emerging trophozoites in TYI-S-33 medium were done. In vivo, the response was evaluated experimentally through counting oocysts out-put every other day until the infection eradicated from the stools of infected E-64 treated mice compared to untreated. Also, the histopathological examination of the small intestine was compared between both of the infected groups. In the present study G. lamblia cysts incubated with E 64 in vitro completely failed in excystation in 90% while trophozoites released on 10% (partially excysted on 5% and completely excysted on 5%) compared to 90 % completely excysted on other non incubated (without E-64) of cysts beside, the trophozoites didn't release on 10% (partially excysted on 5% & completely non-excysted on 5%). In vivo, the evaluation of the therapeutic response proved that the decreasing in the oocysts out-put counting every other day until the infection eradicated from the stools of infected treated mice was very marked in comparison to untreated mice. The differences were statistically significant. The histopathological examination of the small intestine of infected non treated group proved that all the different pathological grades were found while in infected E-64 treated group, only grade I was detected. So, E-64 showed a good therapeutic effect which raises its use in the treatment of human giardiasis