HMEC-1 cells first described by Ades et al. [20 (link)] were cultured in MCDB 131 medium (Gibco, Grand Island, NY) supplemented with L-glutamine (10 mmol/L; Gibco), mouse epidermal growth factor (10 ng/mL; BD Bioscience, San Jose, CA), hydrocortisone (1 μg/mL; Sigma Aldrich, St. Louis, MO), and 10% heat-inactivated fetal bovine serum. Cells were grown in a humidified incubator with 5% CO2 at 37°C and used at approximately 95% confluence. Cells were inoculated at an MOI of 5 based on real-time PCR data. Intracellular growth occurred in a humidified incubator with 5% CO2 at 34°C until times indicated for each experiment.
Mouse epidermal growth factor
Manufactured by BD
Sourced in United States
Mouse epidermal growth factor is a protein that stimulates the growth and differentiation of cells, particularly those of epithelial origin. It plays a crucial role in cell signaling and proliferation.
Automatically generated - may contain errors
Lab products found in correlation
Hydrocortisone, by Merck Group (3 mentions)
Mcdb 131 medium, by Thermo Fisher Scientific (2 mentions)
Cholera toxin, by Merck Group (2 mentions)
L glutamine, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle s medium dmem, by Thermo Fisher Scientific (1 mentions)
Bovine growth serum bgs, by Thermo Fisher Scientific (1 mentions)
Streptomycin, by Merck Group (1 mentions)
Penicillin, by Merck Group (1 mentions)
Lsrfortessa, by BD (1 mentions)
Streptomycin, by BD (1 mentions)
Penicillin, by BD (1 mentions)
Blasticidine, by InvivoGen (1 mentions)
Mouse recombinant epidermal growth factor, by Thermo Fisher Scientific (1 mentions)
Bovine pituitary extract, by Merck Group (1 mentions)
Penicillin streptomycin solution, by Thermo Fisher Scientific (1 mentions)
Mcdb 131 media, by Merck Group (1 mentions)
Keratinocyte serum free medium, by Thermo Fisher Scientific (1 mentions)
Trypsin, by Thermo Fisher Scientific (1 mentions)
Cacl2, by Merck Group (1 mentions)
Neutral protease, by Roche (1 mentions)
6 protocols using mouse epidermal growth factor
1
Culturing and Inoculating HMEC-1 Cells
2
Culturing and Characterizing HPV-Positive Cells
3
Culturing Cell Lines Expressing FcRn
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The HEK293E cell line (ATCC) was cultured in RPMI-1640 GlutaMAX-containing medium (Sigma-Aldrich) supplemented with 10% fetal calf serum (FCS) 25 U/ml penicillin (Sigma-Aldrich) and 25 μg/ml streptomycin (Sigma-Aldrich). The MDCK cell line stably expressing hFcRn (MDCK-hFcRn) was generated by Drs. Jens Fischer and Alex Haas (Roche Pharma Research and Early Development), and cultured in cultured in DMEM with 15% FCS, 25 U/ml penicillin and 25 μg/ml streptomycin, as well as 100 μg/ml G418 (Sigma-Aldrich) to maintain expression of FcRn. HMEC-1 cells stably expressing HA-FcRn-EGFP (HMEC-1-FcRn57 (link)) were cultured in MCDB131 medium (Gibco) supplemented with 2 mM L-glutamine, 25 μg/ml streptomycin, 25 U/ml penicillin, 10% FCS, 10 ng/ml mouse epidermal growth factor (BD Biosciences) and 1 μg/ml hydrocortisone (Sigma-Aldrich), and 100 μg/ml G418 and 50 μg/ml blasticidine (InvivoGen) to maintain FcRn expression. Percentage of HMEC-1 cells expressing HA-FcRn-EGFP in culture was above 85%, which was verified by visualizing EGFP-positive live cells, stained with the amine-reactive dye ViViD (Invitrogen), in combination with a biotinylated anti-FcRn mAb (ADM3195 (link)) and allophycocyanin-conjugated streptavidin (PhycoLink) using a BD LSRFortessa.
4
Isolation and Culture of Epidermal Stem Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The full-thickness skin of newborn WT mice was obtained and incubated with 0.5% Dispase II solution for 6 h at 4 °C to separate the epidermis and dermis. To prepare epidermal single-cell suspensions, the epidermal samples were immersed in 0.3% trypsin solution for 15 min at 37 °C, washed with RPMI medium (containing 10% FBS) and filtered through a 70-µm sterile strainer. For EpSC culture, the filtered cells were centrifuged at 250 × g for harvesting epidermis cells. The pellet was resuspended and cultured with complete medium containing K‐SFM (Gibco), bovine pituitary extract (20‐30 mg/mL), cholera toxin (1 × 10−10 mol/L; Sigma), mouse recombinant epidermal growth factor (0.1‐0.2 ng/mL; Peprotech), mouse epidermal growth factor (10 ng/mL; BD), streptomycin/penicillin solution (100 IU/L; Gibco) and calcium chloride (0.05 mmol/L). The medium was changed every 48 h. The cells at 2–4 passage were used for identification as well as other experiments.
5
Culturing Human Microvascular Endothelial Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human dermal microvascular endothelial cells (HMEC-1) [36] were obtained from the Centers for Disease Control (Atlanta, GA) and maintained in MCDB-131 Media (Sigma-Aldrich, St. Louis, MO) supplemented with 10% heat-inactivated fetal bovine serum (FBS, Atlanta Biological, Atlanta, GA), mouse epidermal growth factor (10 ng/ml, Becton-Dickenson, Bedford, MA), hydrocortisone (1 µg/ml, Sigma-Aldrich), HEPES (10 mM), and pyruvate (1 mM), an atmosphere of 5% CO2 at 37°C. Medium was changed every 3–4 days and cells were passaged once per week. For experiments cells were grown in either 100 mm culture dishes, or gelatin-coated glass coverslips and used for experiments at 2–3 days post-confluence.
6
Isolation and Characterization of Mouse Epidermal Stem Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mouse EpSCs were acquired from the skin of newborn (day 0–2) mice as previously described [12 (link)]. In brief, the separated mouse skin was incubated in 0.5% neutral protease (Roche, Switzerland) at 4°C overnight. The epidermis was carefully separated from the dermis and digested with trypsin (Gibco, USA). Then, the single isolated cells were suspended in keratinocyte serum-free medium (Invitrogen, USA) complemented with CaCl2 (0.05 mM), cholera toxin (10 M; Sigma, USA), streptomycin/penicillin (100 IU/ml) and mouse epidermal growth factor (10 ng/ml; Becton Dickinson Biosciences, USA) and cultured at 37°C in 5% CO2. The Animal Ethics Committee of the Third Military Medical University approved all protocols involving animals. Identification of the primary cells was achieved using flow cytometry and staining with the following fluorochrome-labelled murine monoclonal antibodies: α6-integrin (CD49f) (Invitrogen, USA), transferrin receptor (CD71) (Becton Dickinson Biosciences, USA) and keratin 14 (K14), keratin 15 (K15) and keratin 10 (K10) (all from Santa Cruz Biotechnology, USA).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!