Advertisement

Cytotechnology

, Volume 65, Issue 5, pp 795–802 | Cite as

Optimized flow cytometry protocol for analysis of surface expression of interleukin-1 receptor types I and II

  • Filipp Filippovich Vasilyev
  • Julia Anatolievna Lopatnikova
  • Sergey Vitalievich SennikovEmail author
Method in Cell Science

Abstract

The biological effects of interleukin (IL)-1 are realized through binding to specific membrane-bound receptors. The efficiency of IL-1 action depends on the number of receptors on the cell. We determined the percentage of cells that express IL-1 receptor type I (IL-1RI) and IL-1 receptor type II (IL-1RII) by flow cytometry using phycoerythrin (PE)-labelled antibodies to the IL-1Rs, and the mean absolute number of membrane-bound IL-1Rs per cell using QuantiBRITE PE calibration beads. We showed that different subpopulations of immunocompetent cells expressed different numbers of molecules of membrane-bound IL-1RI and IL-1RII. We also established that when cells were stimulated with bacterial lipopolysaccharide, there was a significant increase in the number of IL-1RI expressed, and a significant decrease in the mean number of IL-1RII molecules per cell. Determination of the mean number of membrane-bound IL-1R molecules using this protocol enables us to obtain precise and reproducible data that are necessary for full evaluation of expression levels.

Keywords

Flow cytometry Interleukin-1 receptor Quantitative analysis Peripheral blood mononuclear cells 

Notes

Acknowledgments

This study was supported by the Federal Target Program “Scientific and scientific-pedagogical personnel of innovative Russia” for 2009–2013 (state contract no. 02.740.11.0707).

References

  1. Böyum A (1968) Separation of leukocytes from blood and bone marrow. Introduction. Scand J Clin Lab Invest Suppl 97:7Google Scholar
  2. Colotta F, Re F, Muzio M, Bertini R, Polentarutti N, Sironi M, Giri JG, Dower SK, Sims JE, Mantovani A (1993) Interleukin-1 type II receptor: a decoy target for IL-1 that regulated by IL-4. Science 261:472–475CrossRefGoogle Scholar
  3. D’hautcourt JL (2002) Quantitative flow cytometric analysis of membrane antigen expression. Curr Protoc Cytom 6.12.1–6.12.22Google Scholar
  4. Dinarello CA (1991) Interleukin-1 and interleukin-1 antagonism. Blood 77:1627–1652Google Scholar
  5. Dinarello CA (2001) IL-1 receptor type I. In: Oppenheim JJ, Feldmann M, Durum SK, Hirano T, Vilcek J, Nicola NA (eds) Cytokine reference: A compendium of cytokines and other mediators of host defense. Academic Press, London, pp 1587–1600Google Scholar
  6. Dinarello CA (2010) IL-1: discoveries, controversies and future directions. Eur J Immunol 40:599–606CrossRefGoogle Scholar
  7. Donati D, Degiannis D, Mazzola E, Gastaldi L, Raskova J, Raska KJ, Camussi G (1997) Interleukin-1 receptors and receptor antagonist in haemodialysis. Nephrol Dial Transplant 12:111–118CrossRefGoogle Scholar
  8. Dower SK, Kronheim SR, March CJ, Conion PJ, Hopp TP, Gillis S, Urdal DL (1985) Detection and characterization of high affinity plasma membrane receptors for human interleukin 1. J Exp Med 162:501–515CrossRefGoogle Scholar
  9. Gudipaty L, Humphreys BD, Buell G, Dubyak GR (2001) Regulation of P2X(7) nucleotide receptor function in human monocytes by extracellular ions and receptor density. Am J Physiol Cell Physiol 280:C943–C953Google Scholar
  10. Jasper GA, Arun I, Venzon D, Kreitman RJ, Wayne AS, Yuan CM, Marti GE, Stetler-Stevenson M (2011) Variables affecting the quantitation of CD22 in neoplastic B cells. Cytometry B Clin Cytom 80:83–90Google Scholar
  11. Mantovani A, Locati M, Vecchi A, Sozzani S, Allavena P (2001) Decoy receptors: a strategy to regulate inflammatory cytokines and chemokines. Trends Immunol 22:328–336CrossRefGoogle Scholar
  12. Matsushima K, Akahoshi T, Yamada M, Furutani Y, Oppenheim JJ (1986) Properties of a specific interleukin 1 (IL1) receptor on human Epstein Barr virus-transformed B lymphocytes: identity of a receptor for IL 1-alpha and IL 1-beta. J Immunol 136:4496–4502Google Scholar
  13. Moraga I, Harari D, Schreiber G, Uzé G, Pellegrini S (2009) Receptor density is key to the Alpha2/Beta interferon differential activities. Mol Cell Biol 29:4778–4787CrossRefGoogle Scholar
  14. Orlova DY, Borisov VI, Kozhevnikov VS, Maltscev VP, Chernyshev AV (2011) Distribution function approach to the study of the kinetics of IgM antibody binding to FcγRIIIb (CD16b) receptors on neutrophils by flow cytometry. J Theor Biol 290:1–6CrossRefGoogle Scholar
  15. Pannu KK, Joe ET, Iyer SB (2001) Performance evaluation of QuantiBRITE phycoerythrin beads. Cytometry 45:250–258CrossRefGoogle Scholar
  16. Penton-Rol G, Orlando S, Polentarutti N, Bernasconi S, Muzio M, Introna M, Mantovani A (1999) Bacterial lipopolysaccharide causes rapid shedding, followed by inhibition of mRNA expression, of the IL-1 type II receptor, with concomitant up-regulation of the type I receptor and induction of incompletely spliced transcripts. J Immunol 162:2931–2938Google Scholar
  17. Reynes J, Portales P, Segondy M, Baillat V, André P, Réant B, Avinens O, Couderc G, Benkirane M, Clot J, Eliaou JF, Corbeau P (2000) CD4 + T cell surface CCR5 density as a determining factor of virus load in persons infected with human immunodeficiency virus type 1. J Infect Dis 181:927–933CrossRefGoogle Scholar
  18. Rossmann ED, Lenkei R, Lundin J, Mellstedt H, Osterborg A (2007) Performance of calibration standards for antigen quantitation with flow cytometry in chronic lymphocytic leukemia. Cytometry B Clin Cytom 72:450–457Google Scholar
  19. Saccani S, Polentarutti N, Penton-Rol G, Sims JE, Mantovani A (1998) Divergent effects of LPS on expression of IL-1 receptor family members in mononuclear phagocytes in vitro and in vivo. Cytokine 10:773–780CrossRefGoogle Scholar
  20. Sims JE, Gayle MA, Slack JL, Alderson MR, Bird TA, Giri JG, Colotta F, Re F, Mantovani A, Shanebeck K, Grabstein KH, Dower SK (1993) Interleukin 1 signaling occurs exclusively via the type I receptor. Proc Natl Acad Sci USA 90:6155–6159CrossRefGoogle Scholar
  21. Stadnichuk IN (1990) Phycobiliproteins. VINITI, MoscowGoogle Scholar
  22. Stylianou E, O’Neill LA, Rawlinson L, Edbrooke MR, Woo P, Saklatvala J (1992) Interleukin 1 induces NF-kappa B through its type I but not its type II receptor in lymphocytes. J Biol Chem 267:15836–15841Google Scholar
  23. Takeuchi M, Nagai S, Tsutumi T, Mio T, Izumi T (1999) The number of interleukin 1 receptors on lung fibroblasts in patients with idiopathic pulmonary fibrosis. Respiration 66:236–241CrossRefGoogle Scholar
  24. Wang L, Abbasi F, Jasper GA, Kreitman RJ, Liewehr DJ, Marti GE, Stetler-Stevenson M (2011) Variables in the quantification of CD4 in normals and hairy cell leukemia patients. Cytometry B Clin Cytom 80:51–56Google Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Filipp Filippovich Vasilyev
    • 1
  • Julia Anatolievna Lopatnikova
    • 1
  • Sergey Vitalievich Sennikov
    • 1
    Email author
  1. 1.Laboratory of Molecular Immunology, Federal State Budgetary Institution “Research Institute of Clinical Immunology”Russian Academy of Medical Sciences Siberian BranchNovosibirskRussia

Personalised recommendations