Cystic Fibrosis pp 15-33

Part of the Methods in Molecular Biology book series (MIMB, volume 742)

Imaging CFTR Protein Localization in Cultured Cells and Tissues



CFTR functions as a chloride channel at the apical membrane of airway, gastrointestinal, and other epithelial cells. Immunofluorescence microscopy is commonly used to assess the subcellular localization and relative abundance of CFTR. Visualization of heterologously overexpressed CFTR is typically unproblematic and straightforward, whereas detection of small quantities of endogenous CFTR in tissues can be challenging and requires highly specific antibodies and optimized staining protocols. CFTR tagged by green fluorescent protein can be employed to study trafficking in live cells. Tagging of CFTR with an extracellular epitope permits detection exclusively at the cell surface and subsequent chasing allows visualization of endocytic trafficking.

Key words

CFTR immunostaining CFTR expression CFTR localization airway epithelium F508del-CFTR 


  1. 1.
    Kartner, N., Augustinas, O., Jensen, T. J., Naismith, A. L., and Riordan, J. R. (1992) Mislocalization of DF508 CFTR in cystic fibrosis sweat gland. Nat Genet 1, 321–327.PubMedCrossRefGoogle Scholar
  2. 2.
    Mall, M., Kreda, S. M., Mengos, A., Jensen, T. J., Hirtz, S., Seydewitz, H. H., et al. (2004) The DeltaF508 mutation results in loss of CFTR function and mature protein in native human colon. Gastroenterology 126, 32–41.PubMedCrossRefGoogle Scholar
  3. 3.
    Kreda, S. M., Mall, M., Mengos, A., Rochelle, L., Yankaskas, J., Riordan, J. R., et al. (2005) Characterization of wild-type and {Delta}F508 cystic fibrosis transmembrane regulator in human respiratory epithelia. Mol Biol Cell 16, 2154–2167.PubMedCrossRefGoogle Scholar
  4. 4.
    Claass, A., Sommer, M., de, J. H., Kälin, N., and Tümmler, B. (2000) Applicability of different antibodies for immunohistochemical localization of CFTR in sweat glands from healthy controls and from patients with cystic fibrosis. J Histochem Cytochem 48, 831–837.PubMedCrossRefGoogle Scholar
  5. 5.
    Gentzsch, M., and Riordan, J. R. (2001) Localization of sequences within the C-terminal domain of the cystic fibrosis transmembrane conductance regulator which impact maturation and stability. J Biol Chem 276, 1291–1298.PubMedCrossRefGoogle Scholar
  6. 6.
    Gentzsch, M., Chang, X. B., Cui, L., Wu, Y., Ozols, V. V., Choudhury, A., et al. (2004) Endocytic trafficking routes of wild type and DeltaF508 cystic fibrosis transmembrane conductance regulator. Mol Biol Cell 15, 2684–2696.PubMedCrossRefGoogle Scholar
  7. 7.
    Cholon, D. M., O’Neal, W. K., Randell, S. H., Riordan, J. R., and Gentzsch, M. (2010) Modulation of endocytic trafficking and apical stability of CFTR in primary human airway epithelial cultures. Am J Physiol Lung Cell Mol Physiol 298, L304–L314.Google Scholar
  8. 8.
    Fulcher, M. L., Gabriel, S., Burns, K. A., Yankaskas, J. R., and Randell, S. H. (2004) Well-differentiated human airway epithelial cell cultures. Methods Mol Med 107, 183–206.Google Scholar
  9. 9.
    Gentzsch, M., Cui, L., Mengos, A., Chang, X. B., Chen, J. H., and Riordan, J. R. (2003) The PDZ-binding chloride channel ClC-3B localizes to the Golgi and associates with cystic fibrosis transmembrane conductance regulator-interacting PDZ proteins. J Biol Chem 278, 6440–6449.PubMedCrossRefGoogle Scholar
  10. 10.
    Engelhardt, J. F., Yankaskas, J. R., Ernst, S. A., Yang, Y., Marino, C. R., Boucher, R. C., et al. (1992) Submucosal glands are the predominant site of CFTR expression in human bronchus. Nat Genet 2, 240–247.PubMedCrossRefGoogle Scholar
  11. 11.
    Kälin, N., Claass, A., Sommer, M., Puchelle, E., and Tümmler, B. (1999) DeltaF508 CFTR protein expression in tissues from patients with cystic fibrosis. J Clin Invest 103, 1379–1389.PubMedCrossRefGoogle Scholar
  12. 12.
    Pawley, J. B. (2006) Handbook of Biological Confocal Microscopy. Springer, New York, NY.Google Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  1. 1.Cystic Fibrosis/Pulmonary Research and Treatment Center, University of North Carolina at Chapel HillChapel HillUSA
  2. 2.Department of Cell and Developmental BiologyUniversity of North Carolina at Chapel HillChapel HillUSA

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