A new collaborative study by Euroimmun and researchers in Türkiye reports that both ELISA and immunoblot show excellent agreement with the reference standard — indirect immunofluorescence assay (IFA) — for serological staging of Epstein-Barr virus (EBV) infections.
A widespread infection
EBV is a ubiquitous herpesvirus that infects more than 95% of the global population, usually during childhood. It is a major cause of infectious mononucleosis. After initial infection, EBV establishes latency with potential to reactivate under immunosuppression. It has been linked to several malignancies and autoimmune diseases, including Burkitt’s lymphoma, nasopharyngeal carcinoma, multiple sclerosis and systemic lupus erythematosus.
Key antibodies for EBV diagnostics
Diagnosis of EBV infection is based primarily on detection of specific antibodies against different EBV antigens. Viral capsid antigen (VCA) IgM is the earliest marker of primary infection. VCA IgG emerges soon after and persists for life. Early antigen (EA) IgG appears during the acute phase and typically declines within months. Nuclear antigen (EBNA) IgG arises 6 to 8 weeks after infection and serves as a marker of past infection.
Parallel testing of VCA IgG/IgM, EA IgG and EBNA IgG is recommended — for example by the Centers for Disease Control and Prevention — for differentiation of acute from past infections. VCA IgG avidity testing further enhances diagnostic accuracy by discriminating early and later stages of primary infection based on the maturation of the IgG response. This is especially useful when serological patterns appear atypical.
Comparison of three methods
The reference method for EBV serology is the highly sensitive and specific indirect immunofluorescence assay (IFA). To assess the efficiency of ELISA and immunoblot compared to IFA, the study analysed antibody responses in 200 paediatric patients with suspected EBV mononucleosis and 40 controls subjects.
VCA (IgM, IgG), EA (IgG), EBNA (IgG) were determined using the EUROPLUS BIOCHIP Sequence EBV IFA, Euroimmun EBV ELISAs and the EUROLINE Anti-EBV Profile 2. VCA IgG avidity was additionally assessed by both IFA and ELISA. The automated platforms EUROLabWorkstation ELISA and EUROBlotOne were used to process the ELISAs and immunoblots, respectively.
Strong concordance between assays
As expected, IFA analysis, including avidity determination, enabled clear classification of early, late and past stages of EBV infection in nearly all 240 samples. ELISA and immunoblot demonstrated near-perfect agreement with IFA for EBV staging. The accuracy of the ELISA data was boosted by additionally analysing VCA IgG avidity, helping to resolve unclear or atypical antibody profiles.
Complementary EBV DNA analysis demonstrated a sensitivity of 60%. The authors note that that EBV DNA testing can be useful when serologic results are inconclusive, but it is insufficient on its own for accurately distinguishing acute infection.
Implications for routine diagnostics
The findings highlight that ELISA, with avidity testing, provides a reliable and efficient alternative to IFA for routine EBV diagnostics. As ELISA requires less specialised equipment, expertise and cost than IFA, it may offer a practical solution for laboratories seeking accurate serological staging without the demands of the reference method.
Read the full study
Güvenç et al. Efficiency of Different Serological Methods for Diagnosing Epstein−Barr Virus Infection and Atypical Serological Profiles. Journal of Medical Virology, 2025; 97:e70724 1 of 10 https://doi.org/10.1002/jmv.70724
