Primary human nasal epithelial cells (HNEpC) were purchased from PromoCell (PromoCell, Heidelberg, Germany) and grown in airway epithelial growth medium (PromoCell) according to the manufacturer’s instructions. Primary cotton rat nasal epithelial cells (CRNEC) were isolated from cotton rat nares and cultivated in Dulbecco’s modified Eagle’s medium-F12 (DMEM-F12) (Gibco-BRL) supplemented with 100 U/ml penicillin, 100 µg/ml streptomycin, 10% heat-inactivated fetal bovine serum, and 2 mM glutamine as previously described (24 (link)). A549 lung epithelial cells (American Type Culture Collection [ATCC CCL-185]) were purchased from the ATCC and grown in DMEM-F12 medium (Gibco-BRL) supplemented with 100 U/ml penicillin, 100 µg/ml streptomycin, 10% heat-inactivated fetal bovine serum, and 2 mM glutamine. Epithelial cells were grown at 37°C under 5% CO2.
Airway epithelial growth medium
Manufactured by PromoCell
Sourced in Germany
Airway Epithelial Growth Medium is a cell culture medium designed to support the growth and maintenance of human airway epithelial cells. It provides the necessary nutrients and growth factors for the optimal proliferation and differentiation of these cells in vitro.
Automatically generated - may contain errors
Lab products found in correlation
Purecol, by Advanced BioMatrix (3 mentions)
Transwell insert, by Corning (2 mentions)
Pneumacult ex plus, by STEMCELL (2 mentions)
Pnuemacult ali, by STEMCELL (2 mentions)
Pneumacult ali, by Advanced BioMatrix (2 mentions)
Dmem f12, by Thermo Fisher Scientific (1 mentions)
Hnepc, by PromoCell (1 mentions)
Atcc ccl 185, by American Type Culture Collection (1 mentions)
Dmem f12 medium, by Thermo Fisher Scientific (1 mentions)
25 cm2 flask, by Corning (1 mentions)
75 cm2 flask, by Corning (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Primocin, by InvivoGen (1 mentions)
Hpmec, by PromoCell (1 mentions)
Hbepc, by PromoCell (1 mentions)
Hsaepc, by PromoCell (1 mentions)
Hpaepic, by ScienCell (1 mentions)
Alveolar epithelial cell medium, by ScienCell (1 mentions)
Endothelial cell growth medium mv2, by PromoCell (1 mentions)
5 protocols using airway epithelial growth medium
1
Cultivation of Nasal and Lung Epithelial Cells
2
Isolation and Expansion of Primary Bronchial Epithelial Cells
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Freshly collected bronchial brushings were agitated in media to detach cells and centrifuged (300 × g, 5 min). The resultant cell pellet was resuspended in serum-free, Airway Epithelial Growth Medium (PromoCell) containing manufacturer’s supplements and Primocin (InvivoGen, #ant-pm-1), added to protect against microbial contamination. The cell suspension was seeded in a 25 cm2 flask (Corning) pre-coated with purified type 1 bovine collagen (PureCol, Advanced BioMatrix, #5005). Cells were cultivated at 37 °C in a humidified atmosphere of 5% CO2 and media was refreshed every 48-h. PBECs were passaged at 80% confluence and cryopreserved in PromoCell media with 10% DMSO (Sigma-Aldrich) and 10% FBS (Gibco) (passage #2). When required, PBECs were thawed quickly in a 37 °C water bath and cultured in a pre-coated 75 cm2 flask (Corning) until 80% confluent, then passaged for use in experiments (Supplementary Table S3 ).
3
Human Bronchial Epithelial Cell Cultures
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Primary HBECs were either a gift from Dr. Steve Brody (Wash U) or isolated from explant donor lungs as previously described56 (link) and cultured for one passage in Airway Epithelial Growth Medium (Promocell C-21160). Continued passaging of HBECs was performed in Pneumacult-Ex Plus (StemCell Technologies #05040) and differentiation of cells was performed in differentiation media, either PnuemaCult-ALI (StemCell Technologies #05001) or VALI media57 (link) on Transwell inserts (Corning #3470, #3460). Expansion and differentiation of HBECs was performed according to the Pneumacult-ALI manufacturer guidelines, with the exception that all cell culture surfaces were coated in PureCol (Advanced Biomatrix #5005) according to product guidelines. The use of de-identified HBECs was approved by the IRB of both the University of Southern California and the University of Iowa as “not Human Subjects Research.”
4
Characterization of Primary Human Pulmonary Cell Types
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Primary human pulmonary microvascular endothelial cells (HPMEC), human small airway epithelial cells (HSAEpC), and human bronchial epithelial cells (HBEpC) were obtained from PromoCell (Germany). Human pulmonary alveolar epithelial cells (HPAEpiC) were purchased from ScienCell (USA). The cells were maintained in Endothelial Cell Growth Medium MV2, Small Airway Epithelial Growth Medium, Airway Epithelial Growth Medium (all from PromoCell), and Alveolar Epithelial Cell Medium (ScienCell). Only cells from passages two to six were used. Cells derived from two different donors were analyzed for every cell type. Each individual is indicated by a three digit numeral identity code: HPMEC: donor #011 and #016; HBEpC: donor #359 and #404; HSAEpC: donor #210 und #306; HPAEpiC: donor #144 and #167. Cells were seeded at 10,000 cells/cm2. Vero E6 cells were maintained in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal calf serum.
5
Primary Human Bronchial Epithelial Cell Culture and Differentiation
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Primary HBECs were either a gift from Dr. Steve Brody (Wash U) or isolated from explant donor lungs as previously described [57 (link)] and cultured for one passage in Airway Epithelial Growth Medium (Promocell C-21160). Continued passaging of HBECs was performed in Pneumacult-Ex Plus (StemCell Technologies #05040) and differentiation of cells was performed in differentiation media, either PnuemaCult-ALI (StemCell Technologies #05001) or VALI media [58 (link)] on Transwell inserts (Corning #3470, #3460). Expansion and differentiation of HBECs was performed according to the Pneumacult-ALI manufacturer guidelines, with the exception that all cell culture surfaces were coated in PureCol (Advanced Biomatrix #5005) according to product guidelines. The use of de-identified HBECs was approved by the IRB of both the University of Southern California and the University of Iowa as “not Human Subjects Research.”
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