Abstract: Objective To analyze the clinical value of anti-EBV capsid antigen IgM antibody(VCA-IgM), EBV DNA and peripheral blood atypical lymphocyte in diagnosing infectious mononucleosis(IM) in infants. Methods Total 32 children with IM(observation group) and 33 children with non-IM(control group) in the First Affiliated Hospital of USTC from October 2017 to March 2019 were retrospectively analyzed. ELISA, quantitative PCR and smear microscopy of peripheral blood anticoagulant samples were used to detect the VCA-IgM, serum EBV DNA and the percentage of atypical lymphocyte in the whole blood, and the differences between the two groups were compared. The diagnostic value of each index and the combined detection of the three indicators were evaluated by ROC curve. Results The positive rates of VCA-IgM, EBV DNA and atypical lymphocytes in the observation group were significantly higher than those in the control group(P<0.05). The sensitivity of combined detection of the three indicators(97.43%) was significantly higher than that of the three indexes alone. The area under the curve(AUC) of combined detection of the three indicators(0.975) for diagnosing IM was higher than that of EBV CA-IgM(0.723), EBV DNA(0.928) and atypical lymphocytes(0.961). Conclusion The detection of EBV VCA-IgM, EBV DNA and atypical lymphocytes can be used as important indicators for the diagnosis of IM in children, and the combined detection can significantly improve the sensitivity. It is helpful for the early diagnosis of IM and reduce the misdiagnosis rate.