Mast Cells pp 217-230 | Cite as

Measurement of Mast Cell Cytokine Release by Multiplex Assay

  • Kevin F. Breuel
  • W. Keith De Ponti
Part of the Methods in Molecular Biology book series (MIMB, volume 315)


Mast cells are highly responsive cells that are capable of secreting a variety of inflammatory mediators, including histamine, heparin, serine proteases, leukotrienes, prostaglandins, and thromboxanes. Studies from several laboratories have demonstrated that mast cells have the capacity to produce a variety of cytokines in response to various stimuli. Characterization of the cytokine profiles in mast cells has routinely been determined by the performance of individual enzyme-linked immunosorbent assays. This process is expensive, time-consuming, and requires a great deal of material to characterize multiple cytokines. In this chapter, we describe a multiplex cytokine assay to detect 17 cytokines simultaneously in 50 µL of culture supernatant derived from stimulated human cord blood-derived mast cells.

Key Words

Mast cells cord blood-derived mast cells Luminex® xMAP™ Bio-Plex 17-plex cytokine assay cytokines microsphere multiplex 


  1. 1.
    Abraham, S. N. and Malaviya, R. (1997) Mast cells in infection and immunity. Infect. Immun. 65, 3501–3508.PubMedGoogle Scholar
  2. 2.
    Barrett, K. E. (1992) Effect of histamine and other mast cell mediators on T84 epithelial cells. Ann. N. Y. Acad. Sci. 664, 222–231.CrossRefPubMedGoogle Scholar
  3. 3.
    Cairns, J. A. and Walls, A. F. (1996) Mast cell tryptase is a mitogen for epithelial cells. Stimulation of IL-8 production and intercellular adhesion molecule-1 expression. J. Immunol. 156, 275–283.PubMedGoogle Scholar
  4. 4.
    Galli, S. J. (1993) New concepts about the mast cell. N. Engl. J. Med. 328, 257–265.CrossRefPubMedGoogle Scholar
  5. 5.
    Hebda, P. A., Collins, M. A., and Tharp, M. D. (1993) Mast cell and myofibroblast in wound healing. Dermatol. Clin. 11, 685–696.PubMedGoogle Scholar
  6. 6.
    Malaviya, R., Malaviya, R., and Jakschik, B. A. (1993) Reversible translocation of 5-lipoxygenase in mast cells upon IgE/antigen stimulation. J. Biol. Chem. 268, 4939–4944.PubMedGoogle Scholar
  7. 7.
    Sugimoto, K., Kasuga, F., and Kumagai, S. (1994) Effects of B subunit of cholera toxin on histamine release from rat peritoneal mast cells. Int. Arch. Allergy Immunol. 105, 195–197.CrossRefPubMedGoogle Scholar
  8. 8.
    Kelley, J. L., Chi, D. S., Abou-Auda, W., Smith, J. K., and Krishnaswamy, G. (2000) The molecular role of mast cells in atherosclerotic cardiovascular disease. Mol. Med. Today 6, 304–308.CrossRefPubMedGoogle Scholar
  9. 9.
    Krishnaswamy, G., Kelley, J., Johnson, D., et al. (2001) The human mast cell: functions in physiology and disease. Front Biosci. 6, D1109–D1127.CrossRefPubMedGoogle Scholar
  10. 10.
    Krishnaswamy, G., Hall, K., Youngberg, G., et al. (2002) Regulation of eosinophilactive cytokine production from human cord blood-derived mast cells. J. Interferon Cytokine Res. 22, 379–388.CrossRefPubMedGoogle Scholar
  11. 11.
    Shakoory, B., Fitzgerald, S. M., Lee, S. A., Chi, D. S., and Krishnaswamy, G. (2004) The role of human mast cell-derived cytokines in eosinophil biology. J. Interferon Cytokine Res. 24, 271–281.CrossRefPubMedGoogle Scholar
  12. 12.
    Hahn, S. and Moroni, C. (1994) Modulation of cytokine expression in PB-3c mastocytes by IBMX and PMA. Lymphokine Cytokine Res. 13, 247–252.PubMedGoogle Scholar
  13. 13.
    Bradding, P., Roberts, J. A., Britten, K. M., et al. (1994) Interleukin-4,-5, and-6 and tumor necrosis factor-alpha in normal and asthmatic airways: evidence for the human mast cell as a source of these cytokines. Am. J. Respir. Cell Mol. Biol. 10, 471–480.PubMedGoogle Scholar
  14. 14.
    Krishnaswamy, G., Lakshman, T., Miller, A. R., et al. (1997) Multifunctional cytokine expression by human mast cells: regulation by T cell membrane contact and glucocorticoids. J. Interferon Cytokine Res. 17, 167–176.CrossRefPubMedGoogle Scholar
  15. 15.
    Church, M. K. and Levi-Schaffer, F. (1997) The human mast cell. J. Allergy Clin. Immunol. 99, 155–160.CrossRefPubMedGoogle Scholar
  16. 16.
    Kellar, K. L. and Douglass, J. P. (2003) Multiplexed microsphere-based flow cytometric immunoassays for human cytokines. J. Immunol. Methods 279, 277–285.CrossRefPubMedGoogle Scholar
  17. 17.
    Prabhakar, U., Eirikis, E., and Davis, H. M. (2002) Simultaneous quantification of proinflammatory cytokines in human plasma using the LabMAP assay. J. Immunol. Methods 260, 207–218.CrossRefPubMedGoogle Scholar
  18. 18.
    Carson, R. T. and Vignali, D. A. (1999) Simultaneous quantitation of 15 cytokines using a multiplexed flow cytometric assay. J. Immunol. Methods 227, 41–52.CrossRefPubMedGoogle Scholar
  19. 19.
    de Jager, W., te Velthuis H., Prakken, B. J., Kuis, W., and Rijkers, G. T. (2003) Simultaneous detection of 15 human cytokines in a single sample of stimulated peripheral blood mononuclear cells. Clin. Diagn. Lab Immunol. 10, 133–139.PubMedGoogle Scholar
  20. 20.
    Martins, T. B., Pasi, B. M., Pickering, J. W., Jaskowski, T. D., Litwin, C. M., and Hill, H. R. (2002) Determination of cytokine responses using a multiplexed fluorescent microsphere immunoassay. Am. J. Clin. Pathol. 118, 346–353.CrossRefPubMedGoogle Scholar
  21. 21.
    Saito, H., Ebisawa, M., Tachimoto, H., et al. (1996) Selective growth of human mast cells induced by Steel factor, IL-6, and prostaglandin E2 from cord blood mononuclear cells. J. Immunol. 157, 343–350.PubMedGoogle Scholar
  22. 22.
    Bio-Rad Laboratories. Bio-Plex Cytokine Assay Instruction Manual. Rev D. 7-1-2003. Bio-Rad Laboratories, Inc., Richmond, CA.Google Scholar

Copyright information

© Humana Press Inc. 2006

Authors and Affiliations

  • Kevin F. Breuel
    • 1
  • W. Keith De Ponti
    • 1
  1. 1.Department of OB/GYN, Quillen College of MedicineEast Tennessee State UniversityJohnson City

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