Mouse melanoma B16F10 cells (Korean Cell Line Bank, Seoul, Republic of Korea) were cultured in DMEM medium (Corning, Manassas, VA, USA), supplemented with 10% (v/v) fetal bovine serum and 1% penicillin/streptomycin, in a humidified atmosphere containing 5% CO2 at 37 °C.
B16f10 cells
Manufactured by Korean Cell Line Bank
Sourced in United States
B16F10 cells are a mouse melanoma cell line. They are commonly used in cancer research and drug discovery studies.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Corning (2 mentions)
Streptomycin, by Corning (2 mentions)
Penicillin, by Corning (2 mentions)
Ht1080 cells, by Korean Cell Line Bank (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
α msh, by Merck Group (2 mentions)
Dmem medium, by Corning (1 mentions)
Rpmi 1640, by Corning (1 mentions)
Endothelial cell growth medium 2 egm2, by PromoCell (1 mentions)
Pc 3 cells, by Korean Cell Line Bank (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
P0781 100ml, by Merck Group (1 mentions)
Mda mb 231, by Korean Cell Line Bank (1 mentions)
Du145, by Korean Cell Line Bank (1 mentions)
Egm 2, by PromoCell (1 mentions)
T stim with con a, by BD (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
13 protocols using b16f10 cells
1
Culturing B16F10 Mouse Melanoma Cells
2
Cell Culture Protocols for Various Cell Lines
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HT1080 cells (KCLB No. 10121, human fibrosarcoma), PC-3 cells (KCLB No. 21435, human prostate adenocarcinoma), and B16F10 cells (KCLB No. 80008, murine melanoma) were obtained from Korean Cell Line Bank (Seoul, Korea) in 2012, where they were characterized by DNA fingerprinting. Cells were maintained in RPMI1640 or Dulbecco's modified Eagle's medium (DMEM, Cellgro, Manassas, VA) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FBS; Cellgro) and 100 U/ml penicillin/100 μg/ml streptomycin (Cellgro) at 37°C in a humidified 5% CO2 incubator. Human umbilical vein endothelial cells (HUVECs) purchased from Innopharmascreen (Asan, Korea) in October 2012 were maintained in Endothelial cell Growth Medium-2 (EGM-2; PromoCell, Heidelberg, Germany) and used at passages 3 to 8. Cells were routinely screened for Mycoplasma contamination.
3
Photodynamic Therapy on B16F10 Cells
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B16F10 cells were purchased from Korean Cell Line Bank (KCLB, Seoul, Republic of Korea). They were grown in DMEM supplemented with 10% fetal bovine serum (life technologies corporation, Waltham, MA, USA) and 1% Penicillin & Streptomycin (life technologies corporation, Waltham, MA, USA). These cells were cultured at 37 °C in a humidified atmosphere containing 5% CO2. B16F10 cells were plated at a density of 5 × 103 cells per well in 96-well plates and then incubated for 3 h. After treatment of the cells with various concentrations of Ce6, the cells were or were not exposed to irradiation of a 660 nm laser with power densities of 1 J/cm2 and were further incubated for 72 h. An MTT assay was carried out to determine the cell viability at 590 nm by using a microplate reader.
4
Peptide Inhibits Tyrosinase Activity
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Example 4
We evaluated whether the peptide fragment of the present invention inhibits tyrosinase activity. B16F10 cells (Korean Cell Line Bank, Seoul) were added to each well of a 24-well plate (5×104 cells per well), along with 1 ml of DMEM, and then stabilized through incubating in a 5% CO2 incubator at 37° C. for 24 hours. α-MSH (Sigma Co, MO, USA), a melanocyte-stimulating hormone, was treated to each well, along with the peptide of SEQ ID NO: 8 in predetermined concentrations. After incubating the cells for 72 hours, the proteins were extracted therefrom. The obtained extracts were added to each well of a 96-well plate (30 g per each well) and then treated with a L-dihydroxyphenylalanine (L-DOPA) solution (100 μL) at 37° C. for 2 hours. The groups in which α-MSH (100 nM) and arbutin (0.01%) were respectively treated were used as positive controls. After 2 hours, the pictures thereof were taken and the tyrosinase activities thereof were measured at 490 nm with a microreader.
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5
Peptide Inhibition of Melanin Production
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Example 3
B16F10 cells (Korean Cell Line Bank, Seoul) were added to each well of a 6-well microplate (1.5×105 cells per well), along with 2 ml of DMEM, and then incubated in a 5% CO2 incubator at 37° C. for 24 hours. α-MSH (Sigma Co, MO, USA) (100 nM) was treated to each well, along with the peptide of SEQ ID NO: 8 in the final concentrations of 0.1, 1, and 10 μM. The groups in which α-MSH (100 nM) and arbutin (0.01%) were respectively treated were used as positive controls. After additionally incubating the cells for 24 hours, the pictures of each culture ware taken so as to compare the respective levels of melanin formation.
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6
Murine Melanoma B16F10 and HUVEC Culture
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Murine melanoma B16F10 cells were purchased from the Korean Cell line Bank (80008, Seoul, Korea) and cultured in Dulbecco's modified Eagle's medium (DMEM) (11995065, Gibco, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS) (EF-0500-A, Atlas Biologicals, For Collins, CO, USA), 100 U/mL penicillin, and 100 μg/mL streptomycin (P0781-100 ML, Sigma-Aldrich, St. Louis, MO, USA). Human umbilical vein endothelial cells (HUVECs) and their growth medium MV2 were purchased from Promo Cell (C-22010, Heidelberg, Germany). The cells were used at passages 4-5 in all experiments. B16F10 cells and HUVECs were incubated at 37°C with 5% CO2. Methyl gallate (MG, PHL82592, Sigma-Aldrich) was dissolved in dimethyl sulfoxide (DMSO) to obtain a stock solution at 500 mM (for in vitro use) or 4 mg/ml (for in vivo use), which was then diluted with autoclaved PBS buffer.
7
Murine Melanoma Cell Viability Assay
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Murine melanoma B16F10 cells were purchased from the Korean Cell Line Bank (KCLB, Seoul, Korea) and cultured in DMEM supplemented with 10% FBS and 1% streptomycin, and then incubated at 37 °C, humidified with 5% atmospheric CO2. Cell viability analyses were performed as previously described [42] (link). Briefly, cells seeded at a density of 1 × 104 cells/well in a 96-well plate for 24 h. On the following day, the cells were exposed to different concentrations of BID3 and incubated for 24 or 48 h, respectively. Then, 10 µL EZ-Cytox solution was added to each well and the cells were incubated for 2–4 h. Absorbance was determined using ELISA at a wavelength of 450 nm. Each assay was performed in triplicate.
8
Cell Culture Protocols for Cancer Research
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Human fibrosarcoma HT1080 cells (KCLB No. 10121), human breast adenocarcinoma MDA-MB231 (KCLB No. 30026), human prostate carcinoma DU145 (KCLB No. 30081), and murine melanoma B16F10 cells (KCLB No. 80008) were purchased from Korean Cell Line Bank (Seoul, Korea). Cells were grown in Roswell Park Memorial Institute (RPMI) 1640 or Dulbecco’s modified Eagle’s medium (DMEM; Lonza, Walkersville, MD, USA) supplemented with 10% FBS (Cellgro, Manassas, VA, USA) and antibiotics (100 U/ml penicillin/100 μg/ml streptomycin; Cellgro) at 37 °C in a humidified 5% CO2 incubator. HUVECs obtained from Innopharmascreen (Asan, Korea) were maintained in EGM-2 (PromoCell, Heidelberg, Germany) and used for experiments at passages 3 to 8.
9
Culturing B16F10 and CTLL-2 Cell Lines
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B16F10 cells (Korean Cell Line Bank, Seoul, Republic of Korea) and CTLL-2 cells (American Type Culture Collection, Manassas, VA, USA) were cultured according to the manufacturer’s instructions. B16F10 cells were cultured in Dulbecco’s Modified Eagle’s Medium with 10% fetal bovine serum. CTLL-2 cells were cultured in RPMI 1640 medium containing 10% T-cell culture supplement with Con A (T-STIM with Con A, BD Biosciences, San Jose, CA, USA) and 10% fetal bovine serum.
10
Generation of Tumor-Conditioned Media
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Supernatant obtained from B16F10 cells (Korean Cell Line Bank) was used to mimic the tumor microenvironment [3 (link)]. B16F10 cells were seeded and cultured with complete DMEM in a 100-mm cell culture dish at 37°C and 5% CO2 until confluency (~90%). The supernatant was collected and then filtered through a 0.2-μm filter. The filtered-supernatant was aliquoted and stored at -20°C until used. We named the B16F10-conditioned media “100% TCM” which was regarded as glucose-depleted because these cells potentially consumed nearly all the glucose in complete DMEM while growing. Two types of TCM mixture (% of TCM) composed of B16F10-conditioned media and DMEM were used; one was “glucose-containing TCM mixture” that consisted of 100% TCM and 25 mM glucose-containing complete DMEM, and the other was a “glucose-depleted TCM mixture” that was composed of 100% TCM and glucose-free DMEM.
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