Optimization of the cytokine secretion assay for human IL-2 in single and combination assays.

ABSTRACT: The cytokine secretion assay identifies live cytokine-secreting cells by capturing the secreted cytokine on a surface-bound capture antibody in dilute suspension culture, followed by detection with a fluorescent anti-cytokine antibody. However, examining the kinetics of cytokine detection revealed that IL-2 staining reached a maximum at early times and then declined, whereas staining for other cytokines including interferon (IFN?) increased for up to 90 min. The decline in IL-2 staining could have been due to rapid cessation of cytokine synthesis, coupled with internalization of cytokine/antibody complexes from the cell surface. Consistent with this model, addition of the anti-IL-2 detection antibody during the cytokine secretion step resulted in higher and more sustained staining. This modified method enhanced staining of IL-2 and IL-4, but not IFN?, tumor necrosis factor alpha (TNF?), or IL-5. However, the longer secretion times possible in the modified assay also improved detection of other cytokines in multi-cytokine combinations.

SUBMITTER: Deng N 

PROVIDER: S-EPMC4759652 | biostudies-literature | 2015 Aug

REPOSITORIES: biostudies-literature