In an effort to develop effective anti-amoebic agent, a salt type complex, namely Ce(Pic)2(H2O)(EO4)·H2O complex, was synthesized and screened for in vitro anti-amoebic activity against Acanthamoeba sp. This Ce(III) complex has been characterized by microelemental analysis, and IR and NMR spectroscopy, and the data suggested the structural formula. Single X-ray crystallography structure showed that the Ce(III) ion is bicapped square antiprismatic with the O2 and O5 atoms occupying the top of the capping position forming a 10-coordination number. The cytotoxicity indicated that Ce salt and Ce(III) complex have an excellent anti-amoebic activity with IC50 values as low as 3.75 and 3 µg/mL, respectively, with a significant decrease (p < 0.05) in Acanthamoeba sp. viability. The mode of cell death in Acanthamoeba cells upon treated with the Ce(III) complex was in the early apoptosis and necrosis. This is in contrast to the Ce(NO3)3·6H2O salt which exhibited necrosis. Both Ce(III) complex and Ce salt on the Acanthamoeba cell exhibited the genotoxic effect on the amoeba cells by looking at scoring DNA damage in types 3 and 4. The complexation form of Ce salt, picric acid (HPic) and tetraethylene glycol (EO4) to produce Ce(III) complex is expected to exhibit synergistic effects to show anti-amoebic activity. The molecular formula of Ce(III) complex and crystal packing hydrogen bonding of this compound is responsible for its cytotoxic and genotoxic effects. Overall results suggested the promising cytotoxic and genotoxic activities of monoclinic Ce(III) complex against Acanthamoeba sp., the preparation method using slow evaporation as a synthetic strategy to design lanthanide complex and the advancement in the synthesis of novel lanthanide complexes with comprehensive evaluation as anti-amoebic agents.