A whole blood intracellular cytokine assay optimised for field site studies demonstrates polyfunctionality of CD4+ T cells in acute scrub typhus
Issued Date
2023-03-01
Resource Type
ISSN
19352727
eISSN
19352735
Scopus ID
2-s2.0-85151550853
Pubmed ID
36961865
Journal Title
PLoS Neglected Tropical Diseases
Volume
17
Issue
3
Rights Holder(s)
SCOPUS
Bibliographic Citation
PLoS Neglected Tropical Diseases Vol.17 No.3 (2023)
Suggested Citation
Inthawong M., Pinthong N., Thaiprakhong A., Wangrangsimakul T., Sunyakumthorn P., Hill J., Sonthayanon P., Paris D.H., Dunachie S.J., Kronsteiner B. A whole blood intracellular cytokine assay optimised for field site studies demonstrates polyfunctionality of CD4+ T cells in acute scrub typhus. PLoS Neglected Tropical Diseases Vol.17 No.3 (2023). doi:10.1371/journal.pntd.0010905 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/82668
Title
A whole blood intracellular cytokine assay optimised for field site studies demonstrates polyfunctionality of CD4+ T cells in acute scrub typhus
Other Contributor(s)
Abstract
Background Assessment of cellular immune responses by combining intracellular cytokine staining and immunophenotyping using flow cytometry enables the simultaneous measurement of T cell phenotype and effector function in response to pathogens and vaccines. The use of whole blood samples rather than peripheral blood mononuclear cells avoids both the need for immediate processing and loss of functional antigen presenting cells due to processing and cryopreservation. Using whole blood provides the possibility to stimulate peripheral T cells in situ, and is more suitable for studies where sample volume is limited, such as those involv-ing children, the elderly and critically ill patients. The aim of this study was to provide a robust tool for the assessment of antigen-specific T cell responses in a field site setting with limited resources. Methodology/principle findings We optimised a flow cytometry-based whole blood intracellular cytokine assay (WBA) with respect to duration of antigen stimulation and intracellular protein retention time. We demonstrate the ability of the WBA to capture polyfunctional T cell responses in the context of acute scrub typhus infection, by measuring IFN-γ, TNF and IL-2 in CD4+ and CD8+ T cells in response to the causative agent O. tsutsugamushi (OT). Using an optimised OT antigen preparation, we demonstrate the presence of polyfunctional antigen-specific memory CD4+ T cells in the blood of scrub typhus patients. Conclusions/significance In conclusion, this flow cytometry-based WBA is well-suited for use at field study sites, and enables the assessment of polyfunctional T cell responses to infectious agents and vaccines through delineation of antigen-specific cytokine secretion at the single cell level.