BACKGROUND Although anti-viral capsid antigen (VCA)-immunoglobin M (IgM) is the most reliable serological marker of primary Epstein-Barr virus (EBV) infection, it could only be detected in limited cases of infectious mononucleosis in children. We analyzed anti-EBV antibodies by an enzyme-linked immunosorbent assay (ELISA), a sensitive method for detecting IgM antibody and compared these results with those obtained by a conventional indirect immunofluorescence (IF) method. METHODS Anti-Epstein-Barr virus early antigen (EA)-IgM and nuclear antigen 1 (EBNA1)-IgG were examined by an ELISA assay in 180 sera from 70 infants and children with infectious mononucleosis, diagnosed serologically by standard IF methods. RESULTS Although by IF, VCA-IgM was detected in only 37 of 70 (52.9%) of the sera from the acute phase of the disease, by ELISA, EA-IgM was detected in 65/70 (92.9%) of these sera. Among infants less than 12 months of age. EA-IgM was positive in 11/13 cases (84.6%) while VCA-IgM was detected in only 3/13 cases (23.1%). Anti-Epstein-Barr virus nuclear antigen 1-IgG was not detected by ELISA in the sera from the acute phase of infectious mononucleosis. Anti-EBNA was not detected by IF in about one-third of the sera during 6-8 months after onset of the disease, whereas by ELISA, EBNA1-IgG was detected in 93.0%. Sera that were positive or negative for both EA-IgM and EBNA1-IgG by ELISA were observed in several cases after the patients recovered from the disease. CONCLUSIONS Although serodiagnosis by the combination of ELISA for EA-IgM and EBNAI-IgG was more sensitive than IF methods, especially in the case of infants and young children, several patients during convalescence and recovery might be judged as seronegative or as being in highly reactivated states.