Chapter 36 : Cytokine Measurement by Flow Cytometry

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In the years since intracellular cytokine staining (ICS) was first developed (1), this method has become a standard way to analyze the functions of immune cells by flow cytometry. It has been used with specific antigen stimulation (2, 3) to read out T cell populations responsive to vaccines (4–6), pathogens (7, 8), cancer (9–11), or allergens (12), in both human and animal models. The technique has been combined with MHC-peptide multimer staining (11, 13), including combinatorial tetramer staining that allows the interrogation of many epitope specificities at once (14, 15). Combined assays with tetramers can be done by first staining with tetramer(s), then stimulating with a nonspecific agent such as PMA+ionomycin, which bypasses the tetramer-occupied T cell receptors. ICS has also increasingly been used with detection of CD154 (16, 17) or CD107a (18) induction, as markers of activation and degranulation, respectively. When used with cell surface and other intracellular phenotyping markers, such multiparameter ICS assays are a tool of choice for functional characterization of T cells and other cytokine-producing cell populations. ICS has also successfully been converted to the mass cytometry platform (19) allowing the use of many more simultaneous markers without significant spillover between readouts.

Citation: Maecker H. 2016. Cytokine Measurement by Flow Cytometry, p 338-342. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch36
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Workflow of ICS assays. Steps in the ICS assay, with variations and optional stopping points as shown.

Citation: Maecker H. 2016. Cytokine Measurement by Flow Cytometry, p 338-342. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch36
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Comparison of antigen-specific T cell assay features

Citation: Maecker H. 2016. Cytokine Measurement by Flow Cytometry, p 338-342. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch36

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