Protocol

Cystic Fibrosis

Volume 742 of the series Methods in Molecular Biology pp 285-310

Date:

Primary Epithelial Cell Models for Cystic Fibrosis Research

  • Scott H. RandellAffiliated withCystic Fibrosis/Pulmonary Research and Treatment Center, The University of North Carolina at Chapel Hill Email author 
  • , M. Leslie FulcherAffiliated withCystic Fibrosis/Pulmonary Research and Treatment Center, The University of North Carolina at Chapel Hill
  • , Wanda O’NealAffiliated withCystic Fibrosis/Pulmonary Research and Treatment Center, The University of North Carolina at Chapel Hill
  • , John C. OlsenAffiliated withCystic Fibrosis/Pulmonary Research and Treatment Center, The University of North Carolina at Chapel Hill

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Abstract

When primary human airway epithelial (hAE) cells are grown in vitro on porous supports at an air–liquid interface (ALI), they recapitulate in vivo morphology and key physiologic processes. These cultures are useful for studying respiratory tract biology and diseases and for testing new cystic fibrosis (CF) therapies. This chapter gives protocols enabling creation of well-differentiated primary CF and non-CF airway epithelial cell cultures with non-proprietary reagents. We also discuss the production of retroviral and lentiviral vectors, the derivation of hAE cell lines, reporter gene assays, and the evolving science of gene overexpression and knockdown in ALI hAE cultures.

Key words

Respiratory tract differentiation physiology pathogenesis therapy adenovirus lentivirus