Characterization of Human Coronaviruses on Well-Differentiated Human Airway Epithelial Cell Cultures

  • Hulda R. Jonsdottir
  • Ronald DijkmanEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1282)


The human airway serves as the entry point of human respiratory viruses, including human coronaviruses. In this chapter we outline the methods by which we establish fully differentiated airway epithelium and its use for human coronavirus propagation. Additionally, we outline methods for immunofluorescence staining of these cultures for virus detection, characterization of cell tropism, and how to perform antiviral assays and quantify viral replication.

Key words

Human coronavirus Antivirals Cell tropism Human airway epithelial cells Virus detection 



This work was supported by the 3R Research Foundation Switzerland (project 128-11).


  1. 1.
    de Jong PM, van Sterkenburg MA, Hesseling SC et al (1994) Ciliogenesis in human bronchial epithelial cells cultured at the air-liquid interface. Am J Respir Cell Mol Biol 10(3):271–277CrossRefPubMedGoogle Scholar
  2. 2.
    Lin H, Li H, Cho HJ et al (2007) Air-liquid interface (ALI) culture of human bronchial epithelial cell monolayers as an in vitro model for airway drug transport studies. J Pharm Sci 96(2):341–350CrossRefPubMedGoogle Scholar
  3. 3.
    Fuchs S, Hollins AJ, Laue M et al (2003) Differentiation of human alveolar epithelial cells in primary culture: morphological characterization and synthesis of caveolin-1 and surfactant protein-C. Cell Tissue Res 311(1):31–45CrossRefPubMedGoogle Scholar
  4. 4.
    Pyrc K, Sims AC, Dijkman R et al (2010) Culturing the unculturable: human coronavirus HKU1 infects, replicates, and produces progeny virions in human ciliated airway epithelial cell cultures. J Virol 84(21):11255–11263CrossRefPubMedCentralPubMedGoogle Scholar
  5. 5.
    Kindler E, Jonsdottir HR, Muth D et al (2013) Efficient replication of the novel human betacoronavirus EMC on primary human epithelium highlights its zoonotic potential. MBio 4(1):e00611–e00612CrossRefPubMedCentralPubMedGoogle Scholar
  6. 6.
    Thompson CI, Barclay WS, Zambon MC et al (2006) Infection of human airway epithelium by human and avian strains of influenza a virus. J Virol 80(16):8060–8068CrossRefPubMedCentralPubMedGoogle Scholar
  7. 7.
    Bernacki SH, Nelson AL, Abdullah L et al (1999) Mucin gene expression during differentiation of human airway epithelia in vitro. Muc4 and muc5b are strongly induced. Am J Respir Cell Mol Biol 20(4):595–604CrossRefPubMedGoogle Scholar
  8. 8.
    Gray TE, Guzman K, Davis CW et al (1996) Mucociliary differentiation of serially passaged normal human tracheobronchial epithelial cells. Am J Respir Cell Mol Biol 14(1):104–112CrossRefPubMedGoogle Scholar
  9. 9.
    Lechner J, LaVeck M (1985) A serum-free method for culturing normal human bronchial epithelial cells at clonal density. J Tissue Cult Methods 9(2):43–48CrossRefGoogle Scholar
  10. 10.
    Fulcher ML, Gabriel S, Burns KA et al (2005) Well-differentiated human airway epithelial cell cultures. Methods Mol Med 107:183–206PubMedGoogle Scholar
  11. 11.
    Dijkman R, Jebbink MF, Koekkoek SM et al (2013) Isolation and characterization of current human coronavirus strains in primary human epithelial cell cultures reveal differences in target cell tropism. J Virol 87(11):6081–6090CrossRefPubMedCentralPubMedGoogle Scholar
  12. 12.
    Lundin A, Dijkman R, Bergstrom T et al (2014) Targeting membrane-bound viral RNA synthesis reveals potent inhibition of diverse coronaviruses including the middle East respiratory syndrome virus. PLoS Pathog 10(5):e1004166CrossRefPubMedCentralPubMedGoogle Scholar
  13. 13.
    Hamming OJ, Terczynska-Dyla E, Vieyres G et al (2013) Interferon lambda 4 signals via the IFNlambda receptor to regulate antiviral activity against HCV and coronaviruses. EMBO J 32(23):3055–3065CrossRefPubMedCentralPubMedGoogle Scholar
  14. 14.
    Schildgen O, Jebbink MF, de Vries M et al (2006) Identification of cell lines permissive for human coronavirus NL63. J Virol Methods 138(1–2):207–210CrossRefPubMedGoogle Scholar
  15. 15.
    Vijgen L, Keyaerts E, Moes E et al (2005) Development of one-step, real-time, quantitative reverse transcriptase PCR assays for absolute quantitation of human coronaviruses OC43 and 229E. J Clin Microbiol 43(11):5452–5456CrossRefPubMedCentralPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Department of Infectious Diseases and PathobiologyVetsuisse Faculty, University of BernBernSwitzerland
  2. 2.Federal Department of Home AffairsInstitute of Virology and ImmunologyBernSwitzerland

Personalised recommendations