An interferon-gamma release assay for monitoring of T-cell responses to Epstein-Barr virus: Application to multiple sclerosis

BACKGROUND: T-cell responses are important for controlling viral infections, but underutilized in routine diagnostics. In recent years, whole-blood interferon-gamma release assays (IGRAs) have emerged as a simple and reliable method for detecting virus-specific T-cells, making them well suited for clinical use. Monitoring of Epstein-Barr virus (EBV) infection is clinically important in transplant recipients and EBV-associated conditions such as Multiple Sclerosis (MS). However, serological assessment may be confounded by disease-modifying treatments, making T-cell-based assays a robust approach for determining EBV-specific immune status. OBJECTIVE: To develop a whole-blood IGRA for the detection of EBV-specific T-cells and to apply it in a clinical context i.e. in patients with MS (pwMS). METHODS: Blood samples from 50 healthy individuals (HI) and 20 pwMS were stimulated with EBNA1 and one self-designed pool (EIHM). IFNγ levels were measured via ELISA and compared to anti-VCA IgG serostatus. RESULTS: The IGRA reliably detected EBV-specific T-cell responses in HI with prior EBV infection using the EBNA1 and EIHM pools. Anti-VCA IgG-positive HI showed significantly higher IFNγ level than EBV-seronegative HI (p < 0.0001), and these responses correlated with anti-VCA IgG levels (EBNA1: p < 0.0001, rs = 0.54; EIHM: p < 0.0001, rs = 0.72). In pwMS, EBV-specific T-cell responses were detectable before therapy and tended to be higher than in HI. After anti-CD20 treatment, EBV-specific IFNγ responses declined significantly (EBNA1: p = 0.0020; EIHM: p = 0.0078). CONCLUSION: The EBV IGRA reliably detects EBV-specific T-cells in seropositive individuals and enables monitoring of cellular immunity in clinical settings.