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The aim of this study was to identify specific laboratory indices to distinguish Epstein-Barr virus–associated hemophagocytic lymphohistiocytosis (EBV-HLH) in children.In this prospective study, Th1/Th2 cytokines, including IL-2, IL-4, IL-6, IL-10, TNF-α, and IFN-γ, were analyzed in patients with EBV-HLH or sepsis at the onset of disease by flow cytometry.IL-10, IFN-γ, IL-10/IL-6, and IFN-γ/IL-6 were higher and IL-6 was lower in EBV-HLH patients compared to sepsis patient levels. When using the criteria of IL-10 >20.9 pg/mL, IFN-γ >17.9 pg/mL, IL-10/IL-6 >0.5 and IFN-γ/IL-6 >0.7, the sensitivity was 89.8%, 93.2%, 93.2%, and 91.5%, while the specificity was 89.8%, 100%, 94.9%, and 100%, respectively. After treatment of EBV-HLH patients, IL-6, IL-10, TNF-α, and IFN-γ were significantly reduced (IL-6: P < .001; IL-10: P < .001; TNF-α: P = .011; IFN-γ: P < .001).This study showed that IFN-γ, IL-10/IL-6, and IFN-γ/IL-6 are novel specific indicators for differential diagnosis of EBV-HLH. Additionally, IL-6, IL-10, TNF-α, and IFN-γ are useful indices for monitoring the effects of treatment on EBV-HLH.IFN-γ, IL-10/IL-6 and IFN-γ/IL-6 are new, specific indicators for the differential diagnosis of EBV-HLH.IL-6, IL-10, TNF-α, and IFN-γ are useful indices for the observation of a curative effect in EBV-HLH.