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Cytometry B Clin Cytom. 2003 Sep;55(1):52-8.

Comparison of intracellular cytokine production with extracellular cytokine levels using two flow cytometric techniques.

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Department of Immunology, Allergology and Rheumatology, University of Antwerp, Belgium.



We investigated the relation between intracellular cytokine production and extracellular cytokine levels by using two flow cytometric techniques.


A two-color flow cytometric technique was used to measure interleukin (IL)-1beta, IL-6, tumor necrosis factor alpha (TNF-alpha), IL-10, and IL-12 production blocked intracellularly with brefeldin A in lipopolysaccharide (LPS)-stimulated CD14(+) monocytes and IL-2, IL-4, and IFN-gamma production in phorbol-12-mirystate-13-acetate (PMA)-stimulated CD3(+) T lymphocytes in samples from patients with rheumatoid arthritis. A flow cytometric microsphere-based immunoassay was performed to detect cytokine secretion in plasma of PMA- and LPS-stimulated whole blood samples.


There was a strong linear correlation between extracellular quantitative (pg/ml) and intracellular semiquantitative detection of LPS-stimulated IL-1beta, IL-6, IL-10, and IL-12 production (r > 0.9). For lymphocytes, extracellularly detected IL-2 and IFN-gamma correlated well with percentages of cytokine-producing cells (r > 0.8). The percentages of IL-4-positive T cells were moderately correlated with the secreted amounts of IL-4 as detected with the microsphere-based immunoassay (r = 0.7).


Overall, there was a good correlation between semiquantitative intracellular detection of cytokines and the secreted amounts of cytokines detected with the microsphere based immunoassay.

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