IL-12 elispot assays to detect and enumerate IL-12 secreting cells

V Ozenci, M Kouwenhoven, R Press, H Link, Y M Huang

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12 Citations (Scopus)


The cytokine IL-12 promotes Th(1)type immune responses and plays a key role in immune regulation. The complex nature of IL-12 hampered its detection without use of stimulants that might give less relevant information. To detect circulating IL-12 p40, we developed enzyme-linked immunospot (ELISPOT) assays that allow enumeration of IL-12 p40 secreting cells without prior in vitro stimulation of the cells. In parallel, intracellular staining of IL-12 p40 by flow cytometry was performed to compare the two methods. IL-12 p40 secreting cells were detected in healthy subjects at a mean number of 103+/-155 per 10(5)blood mononuclear cells (MNC). Numbers of IL-12 p40 secreting blood MNC correlated with IL-12 p40 positive blood MNC detected by flow cytometry. Bacterial endotoxins and the inflammatory cytokines TNF-alpha and IFN-gamma control IL-12 production by antigen presenting cells. Utilizing IL-12 p40 ELISPOT assays, we could confirm occurrence of elevated numbers of IL-12 p40 secreting blood MNC after stimulation with TNF-alpha, IFN-gamma, LPS, LPS+TNF-alpha or LPS+IFN-gamma, compared to cultures without stimulant. Due to its central role in inflammation and autoimmunity, IL-12 is an attractive target for immunotherapy. IL-12 p40 ELISPOT assays represent a sensitive, specific and reliable tool for investigating the role of IL-12 in both health and disease.

Original languageEnglish
Pages (from-to)1218-24
Number of pages7
JournalCytokine Plus
Issue number8
Publication statusPublished - Aug 2000


  • Adult
  • Aged
  • Enzyme-Linked Immunosorbent Assay/methods
  • Evaluation Studies as Topic
  • Female
  • Flow Cytometry/methods
  • Humans
  • Interferon-gamma/metabolism
  • Interleukin-12/analysis
  • Kinetics
  • Leukocytes, Mononuclear/metabolism
  • Lipopolysaccharides/pharmacology
  • Male
  • Middle Aged
  • Tumor Necrosis Factor-alpha/metabolism

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