Comparison between enzyme-linked immunospot assay and intracellular cytokine flow cytometry assay of cytomegalovirus-specific T-cell response in healthy participants
Issued Date
2026-06-01
Resource Type
eISSN
19326203
Scopus ID
2-s2.0-105040808885
Journal Title
Plos One
Volume
21
Issue
6 June
Rights Holder(s)
SCOPUS
Bibliographic Citation
Plos One Vol.21 No.6 June (2026)
Suggested Citation
Klinmalai C., Apiwattanakul N., Prasongtanakij S., Bruminhent J., Srisala S. Comparison between enzyme-linked immunospot assay and intracellular cytokine flow cytometry assay of cytomegalovirus-specific T-cell response in healthy participants. Plos One Vol.21 No.6 June (2026). doi:10.1371/journal.pone.0349292 Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/117172
Title
Comparison between enzyme-linked immunospot assay and intracellular cytokine flow cytometry assay of cytomegalovirus-specific T-cell response in healthy participants
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Abstract
Human cytomegalovirus (HCMV) usually establishes a lifelong latent infection after primary infection. Reactivation occurs sporadically and is controlled by cell-mediated immune response (CMIR). Monitoring of CMIR against CMV is mandatory in immunocompromised patients to adjust immunosuppressive drugs to prevent serious end-organ damage after CMV reactivation. Intracellular staining (ICS) and enzyme-linked immunospot (ELISPOT) quantifying CMV-specific T-cells are generally used as surrogate markers for CMIR against CMV. Whether the results of these 2 methods correlate well is not known. This study compared the numbers of CMV-specific T-cells identified by ELISPOT and ICS in healthy adult volunteers. Correlation with CMV serological status was explored. Thirty peripheral blood samples from healthy individuals were quantified for IFN-γ-producing cells after stimulation with whole CMV and IE1 using ICS, and IFN-γ-secreting cells after stimulation with whole CMV lysate and IE1 peptide pool using ICS and IFN-γ-secreting cells by ELISPOT. Anti-CMV IgG levels were analyzed concomitantly using a chemiluminescent microparticle immunoassay. There were 30 healthy participants, 15 (50%) male, with a mean age of 37.8 (± 7.6) years. Twenty-eight (93.3%) were seropositive against CMV. The CMV-specific CD3<sup>+</sup> cells, as measured by ICS, were highly correlated with the spot numbers obtained by the ELISPOT, irrespective of CMV antigens used (whole CMV, r=0.7677, p<0.0001; IE1, r=0.6516, p<0.0001). The numbers of CMV-specific CD3<sup>+</sup> cells quantified by IE1 stimulation by these 2 assays were statistically correlated with anti-CMV IgG levels (ICS, r=0.5070, p=0.004; ELISPOT, r=0.4384, p=0.015). Among the 30 participants, CMV-specific T cells were detected in all participants (100%), including the two seronegative individuals. The present study demonstrated that CMV-specific T-cells measured by ICS and ELISPOT assays were well correlated, suggesting that these assays could be used to monitor CMV-specific T-cells. CMV IgG levels may reflect prior CMV infection and CMV-specific cell-mediated immunity (CMIR) in immunocompetent individuals.