Use of a short-term whole blood intracellular staining assay to study the T-cell response in respiratory syncytial virus-infected pediatric patients

A. Frassanito, G. Fedele, P. Leone, R. Nenna, F. Midulla, I. Schiavoni

Article ID: 5279
Vol 32, Issue 5, 2018
DOI: https://doi.org/10.54517/jbrha5279
Received: 8 November 2018; Accepted: 8 November 2018; Available online: 8 November 2018; Issue release: 8 November 2018

Abstract

The aim of the present study was the development of a reliable method to evaluate the pattern of the ongoing T-cell response in young infants affected by respiratory infection. To this purpose, we enrolled 44 infants hospitalized with a diagnosis of respiratory syncytial virus bronchiolitis. After a short-term stimulation of whole blood samples, intracellular IFN-g and IL-4 cytokines were measured in CD4+ and CD8+ T-cell subsets by flow cytometry. A stringent staining and gating strategy was used in order to maximize the reduction of background noise and to exclude false positives. The frequencies of cytokine-producing T-cell subsets, albeit low, were easily quantifiable. Cytokine responses were higher in infants sampled > 7 days from the onset of symptoms. The use of a rigorous strategy for cell staining and gating, coupled with a short-term stimulation of whole blood and a careful evaluation of time elapsed from the onset of symptoms constitutes a convincing approach for future clinical studies.


Keywords

bronchiolitis;respiratory syncytial virus;T-cell;flow-cytometry;cytokines


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