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Isolation and Culture of Human Macrophages

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Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 290))

Abstract

Methods to isolate and culture human monocyte-derived macrophages and alveolar macrophages are described. Monocytes are obtained from buffy-coat preparations by Ficoll density gradient centrifugation, followed by adhesion-mediated purification on tissue culture or gelatin-coated plastic. The monocytes differentiate into macrophages in vitro by culturing in medium containing autologous human fibrin-depleted plasma. Alveolar macrophages can be purified from bronchoalveolar fluid samples by adhesion to tissue culture plastic. If resected lung tissue is available, alveolar macrophages can be obtained by mechanically disrupting the lung parenchyma, followed by adhesion-mediated purification.

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References

  1. Bilyk, N. and Holt, P. G. (1995) Cytokine modulation of the immunosuppressive phenotype of pulmonary alveolar macrophage populations. Immunology 86, 231–237.

    PubMed  CAS  Google Scholar 

  2. Upham, J. W., Strickland, D. H., Bilyk, N., Robinson, B. W., and Holt, P. G. (1995) Alveolar macrophages from humans and rodents selectively inhibit T-cell proliferation but permit T-cell activation and cytokine secretion. Immunology 84, 142–147.

    PubMed  CAS  Google Scholar 

  3. Smith, P. D., Smythies, L. E., Mosteller-Barnum, M., et al. (2001) Intestinal macrophages lack CD14 and CD89 and consequently are down-regulated for LPS-and IgA-mediated activities. J. Immunol. 167, 2651–2656.

    PubMed  CAS  Google Scholar 

  4. Mahoney, J. A., Haworth, R., and Gordon, S. (2000) Monocytes and macrophages, in Haematopoietic and Lymphoid Cell Culture (Dallman, M. J. and Lamb, J. R., eds.), Cambridge University Press, Cambridge, pp. 121–146.

    Google Scholar 

  5. Rosen, H. and Gordon, S. (1987) Monoclonal antibody to the murine type 3 complement receptor inhibits adhesion of myelomonocytic cells in vitro and inflammatory cell recruitment in vivo. J. Exp. Med. 166, 1685–1701.

    Article  PubMed  CAS  Google Scholar 

  6. Fraser, I., Hughes, D., and Gordon, S. (1993) Divalent cation-independent macrophage adhesion inhibited by monoclonal antibody to murine scavenger receptor. Nature 364, 343–346.

    Article  PubMed  CAS  Google Scholar 

  7. Wahl, L. M., Katona, I. M., Wilder, R. L., et al. (1984) Isolation of human mono-nuclear cell subsets by counterflow centrifugal elutriation (CCE). I. Characterization of B-lymphocyte-, T-lymphocyte-, and monocyte-enriched fractions by flow cytometric analysis. Cell Immunol. 85, 373–383.

    Article  PubMed  CAS  Google Scholar 

  8. Faradji, A., Bohbot, A., Schmitt-Goguel, M., et al. (1994) Large scale isolation of human blood monocytes by continuous flow centrifugation leukapheresis and counterflow centrifugation elutriation for adoptive cellular immunotherapy in cancer patients. J. Immunol. Methods 174, 297–309.

    Article  PubMed  CAS  Google Scholar 

  9. Flo, R. W., Naess, A., Lund-Johansen, F., et al. (1991) Negative selection of human monocytes using magnetic particles covered by anti-lymphocyte antibodies. J. Immunol. Methods 137, 89–94.

    Article  PubMed  CAS  Google Scholar 

  10. Dehring, D. J. and Wismar, B. L. (1989) Intravascular macrophages in pulmonary capillaries of humans. Am. Rev. Respir. Dis. 139, 1027–1029.

    PubMed  CAS  Google Scholar 

  11. Brain, J. R. M. and Warner, A. (1997) Pulmonary intravascular macrophages, in Lung Macrophages and Dendritic Cells in Health and Disease (Lipscomb, M. R. S., ed.), Marcel Dekker, New York, Vol. 102, pp. 131–149.

    Google Scholar 

  12. Ettensohn, D. B., Jankowski, M. J., Duncan, P. G., and Lalor, P. A. (1988) Bronchoalveolar lavage in the normal volunteer subject. I. Technical aspects and intersubject variability. Chest 94, 275–280.

    Article  PubMed  CAS  Google Scholar 

  13. Freundlich, B. and Avdalovic, N. (1983) Use of gelatin/plasma coated flasks for isolating human peripheral blood monocytes. J. Immunol. Methods 62, 31–37.

    Article  PubMed  CAS  Google Scholar 

  14. Martinez-Pomares, L., Platt, N., McKnight, A. J., da Silva, R. P., and Gordon, S. (1996) Macrophage membrane molecules: markers of tissue differentiation and heterogeneity. Immunobiology 195, 407–416.

    PubMed  CAS  Google Scholar 

  15. Peiser, L., Gough P. J., Darley, E., and Gordon, S. (2000) Characterisation of macrophage antigens and receptors by immunohistochemistry and fluorescent analysis: expression, endocytosis and phagocytosis, in Macrophages (Paulnock, D. M., ed.), Oxford University Press, Oxford, pp. 61–91.

    Google Scholar 

  16. Brain, J. D. and Frank, R. (1973) Alveolar macrophage adhesion: wash electrolyte composition and free cell yield. J. Appl. Physiol. 34, 75–80.

    PubMed  CAS  Google Scholar 

  17. De Brauwer, E. I., Jacobs, J. A., Nieman, F., Bruggeman, C. A., and Drent, M. (2002) Bronchoalveolar lavage fluid differential cell count. How many cells should be counted? Anal. Quant. Cytol. Histol. 24, 337–341.

    PubMed  Google Scholar 

  18. Kini, S. (2002) Color Atlas of Pulmonary Cytopathology, Springer-Verlag, New York.

    Google Scholar 

  19. The BAL Cooperative Group Steering Committee. (1990) Bronchoalveolar lavage constituents in healthy individuals, idiopathic pulmonary fibrosis, and selected comparison groups. Am. Rev. Respir. Dis. 141, S169–S202.

    Google Scholar 

  20. Kobzik, L. (1997) Methods to study lung macrophages, in Lung Macrophages and Dendritic Cells in Health and Disease (Lipscomb, M. R. S., ed.), Marcel Dekker, New York, Vol. 102, pp. 131–149.

    Google Scholar 

  21. Mason, R., Austyn, J., Brodsky, F., and Gordon, S. (1982) Monoclonal anti-macrophage antibodies: human pulmonary macrophages express HLA-DR (Ia-like) antigens in culture. Am. Rev. Respir. Dis. 125, 586–593.

    PubMed  CAS  Google Scholar 

  22. Helinski, E. H., Bielat, K. L., Ovak, G. M., and Pauly, J. L. (1988) Long-term cultivation of functional human macrophages in Teflon dishes with serum-free media. J. Leukocyte Biol. 44, 111–121.

    PubMed  CAS  Google Scholar 

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© 2005 Humana Press Inc., Totowa, NJ

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Davies, J.Q., Gordon, S. (2005). Isolation and Culture of Human Macrophages. In: Helgason, C.D., Miller, C.L. (eds) Basic Cell Culture Protocols. Methods in Molecular Biology™, vol 290. Humana Press. https://doi.org/10.1385/1-59259-838-2:105

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  • DOI: https://doi.org/10.1385/1-59259-838-2:105

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-284-1

  • Online ISBN: 978-1-59259-838-0

  • eBook Packages: Springer Protocols

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