Human Nasal Epithelial Cells (HNEpCs) (Promocell, Rockville, MD, USA) were cultured on poly-L-lysine coated cover glasses (Matsunami, Osaka, Japan) in Airway Epithelial Cell Growth Medium (Promocell) and maintained in 5% CO2 and 95% air at 37 °C. Twelve hours after stimulation with TDI-HSA, HNEpCs were fixed with 4% sucrose-containing 4% paraformaldehyde (Sigma Chemical Co.) for 20 min. The fixed cells were permeabilized with 0.2% Triton X-100/PBS (Sigma Chemical Co.) for 5 min and blocked with 10% goat serum/PBS for 1 h. Then, an anti-IL-33 antibody (Nessy-1, 1:250, Enzo Life Sciences, Farmingdale, NY, USA) was applied overnight at 4 °C. IL-33 was visualized by isotype-specific secondary antibody conjugated with Alexa 488 (1:200, Molecular Probes, Eugene, OR, USA). A fluorescent microscope (Axio Observer, Carl Zeiss, Oberkochen, Germany) was used to obtain fluorescent images.
Hnepcs
Manufactured by Merck Group
HNEpCs are primary human nasal epithelial cells derived from healthy donors. They are useful for in vitro studies of nasal epithelial cell biology and pharmacology.
Automatically generated - may contain errors
Lab products found in correlation
Hnepc, by PromoCell (2 mentions)
Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (2 mentions)
Nessy 1, by Enzo Life Sciences (1 mentions)
Alexa 488, by Thermo Fisher Scientific (1 mentions)
Axio observer, by Zeiss (1 mentions)
Triton x 100 pbs, by Merck Group (1 mentions)
Airway epithelial cell growth medium, by PromoCell (1 mentions)
Poly l lysine coated cover glasses, by Matsunami (1 mentions)
Rpmi 1640 medium, by PromoCell (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
2 protocols using hnepcs
1
Visualizing IL-33 in Human Nasal Epithelial Cells
2
Modulating miR-155-5p and SIRT1 in HNEpCs
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Primary human nasal epithelial cells (HNEpCs) were obtained from PromoCell GmbH. Cells were grown in a RPMI-1640 medium (cat. no. 31870082, Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (Invitrogen; Thermo Fisher Scientific, Inc.) in a humidified incubator at 37°C with 5% CO2.
HNEpCs were cultured in a 96-well plate (3×105 cells/ml) for 24 h. miR-155-5p inhibitor (5′-ACCCCUAUCACGAUUAGCAUUAA-3′) and inhibitor control (5′-CAGUACUUUUGUGUAGUACAA-3′) were purchased from Sigma-Aldrich, Merck KGaA. The medium in the plates was replaced with a fresh one following culturing for 24 h and the HNEpCs were transfected with 50 nM miR-155-5p inhibitor and inhibitor control using Lipofectamine® 2000 transfection reagent (Thermo Fisher Scientific, Inc.). The cells were harvested at 48 h post-transfection for subsequent studies.
To explore the effect of TGF-β1 on HNEpC morphology and EMT, HNEpCs were stimulated with 5 ng/ml transforming growth factor (TGF)-β1 (cat. no. 34-8348-82; Invitrogen; Thermo Fisher Scientific, Inc.) in the presence of miR-155-5p inhibitor or inhibitor control. HNEpC morphology was observed under a DM500 light microscope (Leica Microsystems, Inc.). Furthermore, to explore the function of SIRT1, an SIRT1 inhibitor, Splitomicin (cat. no. S4068; Sigma-Aldrich; Merck KGaA) at final concentration of 100 µM was incubated with the cells for 48 h at 37°C.
HNEpCs were cultured in a 96-well plate (3×105 cells/ml) for 24 h. miR-155-5p inhibitor (5′-ACCCCUAUCACGAUUAGCAUUAA-3′) and inhibitor control (5′-CAGUACUUUUGUGUAGUACAA-3′) were purchased from Sigma-Aldrich, Merck KGaA. The medium in the plates was replaced with a fresh one following culturing for 24 h and the HNEpCs were transfected with 50 nM miR-155-5p inhibitor and inhibitor control using Lipofectamine® 2000 transfection reagent (Thermo Fisher Scientific, Inc.). The cells were harvested at 48 h post-transfection for subsequent studies.
To explore the effect of TGF-β1 on HNEpC morphology and EMT, HNEpCs were stimulated with 5 ng/ml transforming growth factor (TGF)-β1 (cat. no. 34-8348-82; Invitrogen; Thermo Fisher Scientific, Inc.) in the presence of miR-155-5p inhibitor or inhibitor control. HNEpC morphology was observed under a DM500 light microscope (Leica Microsystems, Inc.). Furthermore, to explore the function of SIRT1, an SIRT1 inhibitor, Splitomicin (cat. no. S4068; Sigma-Aldrich; Merck KGaA) at final concentration of 100 µM was incubated with the cells for 48 h at 37°C.
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