Free-living amoebae belonging to Acanthamoeba genus are widely distributed protozoans which are able to cause infection in humans and other animals such as keratitis and encephalitis. Acanthamoeba keratitis is a vision-threatening corneal infection with currently no available fully effective treatment. Moreover, the available therapeutic options are insufficient and are very toxic to the eye. Therefore, there is an urgent need for the development of more effective anti-amoebic agents. Nanotechnology approaches have been recently reported to be useful for the elucidation antimicrobial, antiviral, antifungal and antiprotozoal activities and thus, they could be a good approach for the development of anti-Acanthamoeba agents. Therefore, this study was aimed to explore the activity and cytotoxicity of tannic acid-modified silver nanoparticles, pure silver nanoparticles and pure gold nanoparticles against clinical strains of Acanthamoeba spp. The obtained results showed a significant anti-amoebic effect of the tannic acid-modified silver nanoparticles which also presented low cytotoxicity. Moreover, tannic acid-modified silver nanoparticles were well absorbed by the trophozoites and did not induce encystation. On the other hand, pure silver nanoparticles were only slightly active against the trophozoite stage and pure gold nanoparticles did not show any activity. In conclusion and based on the observed results, silver nanoparticle conjugation with tannic acid may be considered as potential agent against Acanthamoeba spp.