Entamoeba histolytica was recently reclassified to recognize the existence of two genetically distinct but morphologically indistinguishable species: E. histolytica, the protozoan parasite that causes amebic dysentery and liver abscess, and Entamoeba dispar, a nonpathogenic intestinal parasite. Acceptance of this redefinition has dramatically changed both our understanding of the true epidemiology of E. histolytica and the optimal methods for diagnosing amebiasis. Molecular-based diagnostic tests using polymerase chain reaction (PCR) to amplify amebic DNA or enzyme-linked immunosorbent assay (ELISA) to identify amebic antigens in stool samples have been developed to distinguish infection with E. histolytica from infection with E. dispar. Because of its ability to differentiate strains of E. histolytica, PCR is a very useful research tool. Only the ELISA-based test is simple to perform, making it clinically useful in the developing world. To avoid unnecessary treatment of individuals infected with E. dispar, the World Health Organization has stressed the importance of making a specific diagnosis of E. histolytica infection (not E. histolytica/E. dispar) before treating for amebiasis.