Inflammatory pseudotumors (IPT) are soft tissue tumors that include a diverse group of lesions characterized by inflammatory cell infiltration and variable fibrotic responses. Idiopathic orbital inflammatory pseudotumors (IOIP) are IPTs of unknown etiology that develop in the orbit. Due to the lack of well-defined pathogenic mechanisms, diagnosis and treatment of this disease remain a significant challenge. Epstein-Barr virus (EBV) infection, which causes significant lymphocyte infiltration, has been proposed to be involved in IOIP. This study tries to validate the relationship between EBV infection and the development of IOIP. Sixteen IOIP tissue samples were obtained from patients during surgical resection of the lesion. One Graves' ophthalmopathy tissue sample and 20 normal donors' plasma serves as controls. The plasma level of five EBV antibodies, including VCA-IgG, VCA-IgA, VCA-IgM, EA-IgG and EBNA1-IgG were examined. All plasma samples were EB-VCA-IgG positive and EB-VCA-IgM negative, suggesting that all people tested had been infected with EBV but not in the acute infection stage. EBV-DNA was detected in 15/16 (94%) of IOIP tissue samples despite different levels of lymphocyte infiltration and 5/16 plasma samples (31%) were detected EBV DNA positive which is higher than the normal controls (10%). Percent of positive plus suspected positive samples with one or more of the three important risk markers (VCA-IgA, EA-IgG, EBV-DNA) is 50% of the patients (8/16) which is much higher compare with the normal controls (20%). The results further reveal the relationship between IOIP and EBV infection.