P06 07050

Experiments were performed with two immortalized human airway epithelial cell lines (16HBE14o^-, S9) and one alveolar cancer cell line (A549). With permission of D.C. Gruenert 16HBE14o^- cells were received from K. Kunzelmann (University of Regensburg, Germany). S9 cells were purchased from ATCC-LGC Standards (Wesel, Germany, S9). A549 cells were obtained from the cell collection of the Friedrich Loeffler-Institute (Riems, Germany).
Cells were cultured on 10 cm Cell+ dishes (3902300, Sarstedt, Numbrecht, Germany) in Eagle’s minimal essential medium (MEM) (P03-2950, PAN-Biotech, Aidenbach, Germany) containing 10% FBS superior (S0615, Merck, Darmstadt, Germany), 29.8 mmol/L NaHCO₃ and 1% (w/v) penicillin/streptomycin solution (P06-07050, PAN-Biotech, Aidenbach, Germany) at 37 °C and gassed with 5% CO₂. Cell culture medium of A549 cells additionally contained 1% L-glutamine solution (P04-80100, PAN-Biotech, Aidenbach, Germany). Cell culture medium was changed every 3 days. Just before cells formed confluent monolayers, they were passaged onto new cell culture dishes or used directly for experiments. All cell cultures were checked for Mycoplasma contaminations on a regular basis.

We generated lymphoblastoid cell lines (LCLs) from subjects 1 and 2 from blood samples collected in lithium heparin-coated tubes (BD Vacutainer PST). Red blood cells were lysed through incubation in lysis buffer (155 mM ammonium chloride, 10 mM potassium hydrogen carbonate, and 0.1 mM disodium-EDTA) for 10 min. Pelleted lymphocytes were immortalized by transfection of Epstein-Barr virus, followed by incubation at 37 °C for 1 h. Finally, cyclosporine was added to selectively kill T lymphocytes and generate only a cell line of B lymphocytes as detailed previously.² (link) In short, LCLs were grown in RPMI 1640 Medium, GlutaMAX Supplement (61870044; Thermo Fisher Scientific), supplemented, and fortified with 10% fetal bovine serum (FBS; Biochrom), L-glutamine (P04-80050; PAN Biotech), and antibiotics (penicillin/streptomycin, P06-07050; PAN Biotech). Cells were cultured at 37 °C in incubators supplied with 5% CO₂.