A simple, robust flow cytometry-based whole blood assay for investigating sex differential interferon alpha production by plasmacytoid dendritic cells

• Published in: Journal of immunological methods 2022-05, Vol.504, p.113263-113263, Article 113263
• Main Authors: Sampson, Oliver, Lim, Nicholas, White, Jemima, Vieira, Vinicius, Kløverpris, Henrik, Adland, Emily, Conlon, Chris, Skelly, Donal, Jones, Lucy, Stafford, Lizzie, Jamsen, Anni, Barnes, Ellie, Dunachie, Susie, Frater, John, Klenerman, Paul, Altfeld, Marcus, Goulder, Philip
• Format: Article
• Language: English
• Subjects: Dendritic Cells; Female; Flow Cytometry; Humans; Immune sex differences; Immunity, Innate; Innate immunity; Interferon alpha; Interferon Type I; Male; Plasmacytoid dendritic cell; Toll-like receptor 7
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/j.jim.2022.113263

Central to sex differences observed in outcome from infection and vaccination is the innate immune response, and specifically production of type I interferons by plasmacytoid dendtiric cells (pDCs), the main producers of IFN-α. Evaluation of IFN-α production by pDCs is therefore critical for studies of innate immune function. However, reliable measurement of pDC IFN-α is hampered by reduced cell yields and cytokine production after cryopreservation or after even short delays in stimulating freshly isolated cells. We here describe a simple yet robust method for measuring IFN-α production in pDCs that preserves cell activation and cytokine production through immediate stimulation of whole blood and subsequent maintenance at 37 °C. •Immediate whole blood stimulation detects differential IFN-α production by pDCs•Maintaining whole blood samples at 37 °C preserves maximal cytokine production•6 h incubation best captures differential pDC activation and cytokine production•Assay robustness is confirmed by low intra-donor temporal variability•Cell activation and cytokine production are substantially reduced by cryopreservation