To develop an improved serodiagnostic test for amoebiasis, we performed a detailed analysis of the immunodominant epitopes of the 29 kDa surface antigen and evaluated its sensitivity and specificity. Enzyme-linked immunosorbent assay (ELISA) based on the fragment containing the immunodominant epitope was evaluated further and compared with full-length recombinant 29 kDa protein. Specificity and sensitivity of the two ELISAs were assessed using 55 human sera of parasitic protozoa infection cases (25 amoebiasis, 20 giardiasis and 10 toxoplasmosis sera) and 10 healthy control sera. The immunodominant epitope of the 29 kDa antigen is localised only in the N-terminus 14-54 amino acid residues. The sensitivities of the two ELISAs were very high, 92 and 96%, respectively. The specificity of the fragment was 100%, whereas the specificity of the full-length 29 kDa protein was 86.6%. These results indicate that the fragment containing the immunodominant epitope of the 29 kDa protein can be used to accurately serodiagnose amoebiasis without cross-reactivity from other parasites.