Isolation, cultivation and application of primary respiratory epithelial cells obtained by nasal brushing, polyp sample or lung explants

This protocol provides guidance for the standardised method of performing isolation, maintenance and polarization of the respiratory epithelial primary cells acquired from nasal brushing, polyp specimens or from lung explants. Culture of the airway epithelium is of great importance in the understanding and modelling of different lung diseases including cystic fibrosis (CF).
This method provides a perfectly polarized layer of epithelial cells on filters, with a good number of ciliated cells and a thin layer of mucus. These polarized cells demonstrate ion transport properties thus the filters can potentially be used for electrophysiological measurements using an Ussing system, or for airway surface liquid (ASL) volume analysis or immunofluorescent staining.
This protocol presents all the necessary information for conducting successful cell culture without co-culture of the feeder cells (fibroblasts). It includes two updated procedures for (i) nasal brushing and (ii) freezing/defrosting for biobanking of samples involved in clinical trials within the European Cystic Fibrosis Society - Clinical Trial Network (ECFS-CTN).