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Background: Clinical investigation of antigen-specific T cells in potentially immunodeficient patients is an important and often challenging aspect of patient diagnostic work up. Methods for detection of microbial exposure to the T cell compartment exist but are laborious and time consuming. Recently, a whole blood technique involving flow cytometry and detection of CD25 and OX40 (CD134) expression on the surface of activated CD4+ T cells was shown to be accurate and concordant when compared with more traditional methods of antigen-specific T cell detection. Methods: Whole heparinised blood was collected from healthy donors and set up using the 'OX40' assay to detect antigen specific CD4+ T cell responses to Varicella Zoster Virus (VZV), Epstein-Barr Virus (EBV), Cytomegalovirus (CMV), Candida albicans and Streptococcus pneumoniae. Results: The 'OX40' assay technique was clinically validated for routine use in an NHS clinical immunology laboratory by analysis of incubation length (40-50 hours), sample transport time (up to 24 hours at room temperature), concordance with serology testing, proliferation and IFN-γ production. In addition, 63 healthy controls (age range 21-78) were tested for responses to generate a healthy control reference range. Conclusions: The OX40 assay, as presented in this report, represents an economical, rapid, robust whole blood technique to detect antigen-specific T cells which is suitable for clinical immunology diagnostic laboratory use. © 2013 Clinical Cytometry Society.

Original publication




Journal article


Cytometry B Clin Cytom

Publication Date



CD4+T cells, Co-stimulation/Co-stimulatory, Diagnostics, Flow cytometry