title: Comparison of intracellular cytokine production with extracellular cytokine levels using two flow cytometric techniques.
abstract: BACKGROUND
  We investigated the relation between intracellular cytokine production and extracellular cytokine levels by using two flow cytometric techniques.
  
  
  METHODS
  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.
  
  
  RESULTS
  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).
  
  
  CONCLUSIONS
  Overall, there was a good correlation between semiquantitative intracellular detection of cytokines and the secreted amounts of cytokines detected with the microsphere based immunoassay.
authors.affiliation: Department of Immunology, Allergology and Rheumatology, University of Antwerp, Belgium.
authors.forename: A J
authors.initials: AJ
authors.lastname: Schuerwegh
authors.forename: L S
authors.initials: LS
authors.lastname: De Clerck
authors.forename: C H
authors.initials: CH
authors.lastname: Bridts
authors.forename: W J
authors.initials: WJ
authors.lastname: Stevens
mesh_terms: D001172:Arthritis, Rheumatoid / Q000276:immunology; D017252:CD3 Complex / Q000378:metabolism; D002478:Cells, Cultured; D016207:Cytokines / Q000032:analysis* / Q000187:drug effects; D045604:Extracellular Fluid / Q000737:chemistry*; D005434:Flow Cytometry; D006801:Humans; D007118:Immunoassay; D007424:Intracellular Fluid / Q000737:chemistry*; D018950:Lipopolysaccharide Receptors / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D008863:Microspheres; D009000:Monocytes / Q000737:chemistry / Q000187:drug effects / Q000502:physiology*; D013601:T-Lymphocytes / Q000737:chemistry / Q000187:drug effects / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D017252:CD3 Complex; D016207:Cytokines; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D013755:Tetradecanoylphorbol Acetate