The present study was carried out to investigate the prevalence of fascioliasis among 100 cattle and buffaloes of different age and sex in Assiut Governorate. Samples were examined by microscopic examination, Agar Gel Diffusion Test AGDT and ELISA techniques to detect Fasciola species eggs and antibodies, respectively. Prevalence of Fasciola was 8% by microscopic examination, 32% by AGDT and 60.86% by ELISA test. Cattle showed higher prevalence of infection 61.4% than buffaloes 29.4%. Female cattle and buffaloes were more susceptible to the infection 64.55% than male 23.8%. Fasciola was more frequently recorded in 2-4 years old cattle and buffaloes 76% followed by more than 4 years 46.7% then less than 2 years of age 20%. All positive results with fecal egg examination and AGDT were also positive with ELISA. Serological examination by AGDT and ELISA confirms the microscopical examination. The results of this study revealed that the agar gel diffusion test and ELISA could become a useful tools to diagnose fascioliasis in cattle and buffaloes.