Fasciola gigantica infects buffalo in Asian and African countries and causes significant economic losses. As a kind of zoonotic parasite disease, human infection has been reported and make fascioliasis patients suffer heavy abdominal pain, anemia and workforce loss. Fasciola can modulate host immune responses to survive from immune attack and some mechanisms have been reported with its antigen protein. Protein, as the chief actors within the cell, presents the dynamic changes of body with its specific changes and serves as unexceptionable biomarkers in many disease-detections. But there is not any study about serum proteomic for diagnostic target seeking in buffalo after infection with F. gigantica. Herein, Isobaric Tags for Relative and Absolute Quantitation (iTRAQ) and Parallel Reaction Monitoring (PRM) technologies were used in the present study to reveal the dynamic changes of proteins in buffalo serum after infected with F. gigantica. Six proteins were found significantly regulated in all three groups which reminds us that they could be used as new diagnostic biomarkers in buffalo throughout F. gigantica infection. All these are new findings in buffalo after infection with F. gigantica and could provide a new insight into the potential possibility in this parasite clearance.