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Rotaviruses are the single most important cause of severe acute diarrhea in young children throughout the world. Many studies have shown the important role of rotavirus as a cause of diarrhea in children in both developed and developing countries. Our study aims to determine the prevalence of rotavirus infection among children less than five years old in Ismailia governorate,Egypt and to compare between Enzyme Linked Immunosorbent Assay (ELISA) and Polyacrylamide Gel Electrophoresis (PAGE) as a diagnostic tool for rotavirus infection in acute infantile diarrhea.