Immortalization Method: Cocultured with irradiated 3T3 fibroblast feeder cells in the presence of the RhoA kinase inhibitor Y-27632.
BioSafety Level: II
Organism: Human (H. sapiens)
Species: Human
Source Organs: Airway/Lung
Organ Type: Airway/Lung
Growth Properties: Adherent
Morphology: Epithelial
Markers: MUC5AC/MUC5B
Donor Age: Not disclosed
Propagation:Use of PriCoatTM T25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. Grow cells in ECM-coated culture vessels unless otherwise specified in the Propagation Requirements below.
These cells require inactivated 3T3J2 feeder cells to grow. Plate inactivated 3T3J2 cells on G422-coated culture vessels at a seeding density of 47,000 cells/cm2. Allow cells to attach for 18 - 24 hours before adding Immortalized Human Delta F508 Cystic Fibrosis Cells.
The base medium for this cell line is a 1:3 mixture of Prigrow III medium and Prigrow XII medium available at abm, Cat. No.'s TM003 and TM012. To make the complete growth medium, add the following components to the base medium: fetal bovine serum (TM999)* to a final concentration of 7%, Penicillin/Streptomycin Solution (G255) to a final concentration of 1%, L-glutamine (G275) to a final concentration of 2 mM, hydrocortisone to a final concentration of 25 ng/ml, Recombinant Human EGF (Z100139) to a final concentration of 0.125 ng/ml, Insulin (TM053) to a final concentration of 5 µg/ml, Antibiotic/Antimycotic (100X) (G273) to a final concentration of 1X, Gentamicin (G272) to a final concentration of 10 µg/ml, cholera toxin to a final concentration of 8.6 ng/ml, and Y-27632 to a final concentration of 5 µM.Change media every 2-3 days.Carbon dioxide (CO2): 5%, Temperature: +37.0°C.* Do not use heat-inactivated FBS for cell culture unless specified otherwise.
Quality Control: 1) Whole mount immunofluorescence
Shipping Condition: Dry Ice
Storage Condition: liquid nitrogen or -180°C
Reference: Gentzsch, M., Boyles, S. E., Cheluvaraju, C., Chaudhry, I. G., Quinney, N. L., Cho, C., Dang, H., Liu, X., Schlegel, R., & Randell, S. H. (2017). Pharmacological Rescue of Conditionally Reprogrammed Cystic Fibrosis Bronchial Epithelial Cells. American journal of respiratory cell and molecular biology, 56(5), 568–574. https://doi.org/10.1165/rcmb.2016-0276MA