We passaged the primary NHBE cells (passage 1) three times before differentiating them (passage 4) into a pseudostratified epithelium. For each passage, the cells were grown in a 100-mm culture dish (Corning Inc.) precoated with PurCol (Advanced Biometrics). The cells were maintained in airway epithelial cell (AEC) growth medium (PromoCell) containing AEC supplements (PromoCell), 2% penicillin/streptomycin (Thermos Fisher Scientific), and 1% amphotericin B (Thermos Fisher Scientific) (complete AEC medium) at 37°C in an incubator with 5% CO2. The cells were grown to 90% confluency, and the medium was changed every other day. Confluent monolayers of cells were dissociated with 5 ml of TrypLE (Thermo Fisher Scientific) and pelleted, and one-third of the cells were reseeded in a culture dish containing complete AEC medium for passaging. A549 cells were grown in F-12 medium (Thermo Fisher Scientific) with 10% HyClone fetal bovine serum (GE Healthcare), 2% penicillin/streptomycin, and 1% amphotericin B.
Aec growth medium
Manufactured by PromoCell
AEC growth medium is a specialized cell culture medium designed to support the growth and proliferation of Aortic Endothelial Cells (AECs). It provides the essential nutrients and growth factors required for the in vitro culture of these cells.
Automatically generated - may contain errors
Lab products found in correlation
Penicillin streptomycin, by Thermo Fisher Scientific (6 mentions)
Amphotericin b, by Thermo Fisher Scientific (3 mentions)
Tryple, by Thermo Fisher Scientific (3 mentions)
100 mm culture dish, by Corning (3 mentions)
Aec supplement, by PromoCell (3 mentions)
F12 medium, by Thermo Fisher Scientific (1 mentions)
Hyclone fetal bovine serum, by GE Healthcare (1 mentions)
Amphotericin b, by GE Healthcare (1 mentions)
Penicillin streptomycin, by GE Healthcare (1 mentions)
3 protocols using aec growth medium
1
Differentiation of Primary NHBE Cells
2
Primary NHBE Cell Culture Protocol
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For primary cell culture, we used a previously described protocol (12 (link), 57 (link), 58 (link)). Briefly, NHBE cell monolayer passaging was also performed in a 100-mm culture dish (Corning, Inc.). The culture dish was coated with PureCol (Advanced Biometrics) before seeding cryopreserved NHBE passage 0 (P0) cells, and these cryopreserved cells were thawed in a water bath. The cells were maintained in airway epithelial cell (AEC) growth medium (PromoCell) with AEC supplement (PromoCell), 2% penicillin-streptomycin (Thermo Fisher Scientific), and 1% amphotericin B (Thermo Fisher Scientific) at 37°C in a 5% CO2 incubator. Cells were grown to 90% confluence with a medium change every other day. A confluent monolayer of cells was dissociated with TrypLE (Thermo Fisher Scientific), pelleted, and reseeded into a culture dish containing AEC medium with supplements for passaging. A portion of the cells was stored at −176°C in liquid nitrogen. Cells were passaged up to four times (P4).
3
NHBE Cell Monolayer Passaging Protocol
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We used a previously described protocol (12 (link), 58 (link), 116 (link)). Briefly, NHBE cell monolayer passaging was also performed in a 100 mm culture dish (Corning, Inc.). The culture dish was coated with PureCol (Advanced Biometrics) before seeding cryopreserved NHBE passage zero (P0) cells, and these cryopreserved cells were thawed in a water bath. The cells were maintained in airway epithelial cell (AEC) growth medium (PromoCell) with AEC supplement (PromoCell), 2% penicillin/streptomycin (Thermo Fisher Scientific) and 1% amphotericin B (Thermo Fisher Scientific) at 37 °C in a 5% CO2 incubator. Cells were grown to 90% confluency with a medium change every other day. A confluent monolayer of cells was dissociated with TrypLE (Thermo Fisher Scientific), pelleted, and reseeded into a culture dish containing AEC medium with supplements for passaging. A portion of the cells was stored at −176°C in liquid nitrogen. Cells were passaged up to four times (P4).
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