Long-Term Culture of Distal Airway Epithelial Cells Allows Differentiation Towards Alveolar Epithelial Cells Suited for Influenza Virus Studies

Imai-Matsushima, Aki; Martin-Sancho, Laura; Karlas, Alexander; Imai, Seiichiro; Zoranovic, Tamara; Hocke, Andreas C.; Mollenkopf, Hans-Joachim; Berger, Hilmar; Meyer, Thomas F.

Non-malignant tissue samples were obtained from pneumectomy specimens from the Clinic for Infectious Diseases and Pulmonary Medicine, Charité University Hospital, Berlin under signed informed consent. Scientific usage for experimental purposes was approved by the ethics committee of the Charité University Medicine, Berlin (EA2/079/13). Tissue pieces were processed according to the method by Daum et al. [23] with modifications. Briefly, they were washed with balanced salt solution buffer (BSSB:137 mM NaCl/5.0 mM KCl/0.7 mM Na 2 HPO 4 /10 mM HEPES/5.5 mM glucose/1.2 mM MgSO 4 /1.8 mM CaCl 2 , pH 7.4), minced finely, digested with trypsin (Serva) and elastase (Merck Millipore), passed through a 70 μm filter, centrifuged at 300 g for 5 min, washed twice with BSSB, resuspended in culture medium and plated into flasks previously seeded with irradiated NIH/3 T3-GFP feeders. The cell yield was donor dependent and ranged from 1.0-5.0 × 10 6 cells/g of tissue.