The Epstein-Barr virus (EBV) open reading frame BFRF3 encodes a viral structural capsid protein or tegument protein, VCA p18, that is highly immunogenic in humans. In this study, a cluster of immunodominant epitopes in the C-terminus of VCA p18 has been identified. These epitopes were combined into a single synthetic peptide that was analyzed for diagnostic value in an ELISA. This VCA combined peptide appeared to be an excellent VCA marker with remarkable applications for IgG- and IgM-related EBV diagnostics. The combined peptide reacted with 95% of 159 sera that were IgG VCA-positive by immunofluorescence assay. In addition, 95% of sera from 67 persons with confirmed infectious mononucleosis were positive for IgM antibodies to this VCA combined peptide. An IgG-specific seroconversion could be demonstrated by subsequent serum samples during the acute phase of primary EBV infections. Although 97% of the tested sera from nasopharyngeal carcinoma patients had IgG antibodies reacting positively with the VCA combined peptide, only 61% contained IgA anti-VCA combined peptide.