Quantitative intracellular cytokine measurement: age-related changes in proinflammatory cytokine production

Clin Exp Immunol. 1998 Aug;113(2):213-9. doi: 10.1046/j.1365-2249.1998.00641.x.


The proinflammatory cytokines play a central role in mediating cellular and physiological responses, and levels may reflect immune system effectiveness. In this study, the effect of ageing on the inflammatory response was examined using a novel method to detect production of the proinflammatory cytokines, i.e. tumour necrosis factor-alpha (TNF-alpha), IL-6 and IL-1beta. Peripheral blood mononuclear cells (PBMC) obtained from healthy donors of different ages were incubated for 0, 24, 48 and 72 h with or without phorbol 12-myristate 13-acetate (PMA) stimulation. At each time point these cells were permeabilized and incubated with secondary conjugated FITC MoAbs specific for each cytokine. A flow cytometric system was developed to quantify specific intracellular fluorescence in T cells (CD3+) and monocytes (CD14+). TNF-alpha, IL-6 and IL-1beta production in cell culture supernatants was also measured using ELISAs. In older subjects, flow cytometry detected significant increases in intracellular T cell TNF-alpha and IL-6 (P < 0.05). IL-1beta was not detected in any of the T cell samples. Likewise, the monocytes of older subjects demonstrated increased intracellular levels of all three cytokines, but these increases were not significant (P > 0.05). These changes in intracellular proinflammatory cytokine levels may explain some of the exaggerated inflammatory responses seen in elderly patients.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Aging / immunology*
  • Analysis of Variance
  • CD3 Complex
  • Cytokines / biosynthesis*
  • Enzyme-Linked Immunosorbent Assay
  • Flow Cytometry
  • Humans
  • Interleukin-1 / biosynthesis
  • Interleukin-6 / biosynthesis
  • Leukocytes, Mononuclear / drug effects
  • Leukocytes, Mononuclear / immunology*
  • Lipopolysaccharide Receptors
  • Middle Aged
  • Tetradecanoylphorbol Acetate / pharmacology
  • Tumor Necrosis Factor-alpha / biosynthesis


  • CD3 Complex
  • Cytokines
  • Interleukin-1
  • Interleukin-6
  • Lipopolysaccharide Receptors
  • Tumor Necrosis Factor-alpha
  • Tetradecanoylphorbol Acetate