B16 cells (ATCC Number CRL-6475TM, VA, USA) were maintained in low glucose DMEM (Sigma-Aldrich, Singapore) supplemented with 10% fetal bovine serum (Gibco Life Technologies, Massachusetts, MA, USA), 10 U/mL of penicillin (Gibco Life Technologies, Massachusetts, MA, USA), and 10 mg/mL of streptomycin (Gibco Life Technologies, Massachusetts, MA, USA) in a 37 °C, 5% CO2, and 95% humidified atmosphere.
B16 cells
Manufactured by American Type Culture Collection
Sourced in United States, China
B16 cells are a well-established murine melanoma cell line derived from a C57BL/6 mouse. They are a commonly used model for studying melanoma biology and evaluating anti-cancer therapies.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Dmem low glucose, by Merck Group (1 mentions)
Streptomycin, by Thermo Fisher Scientific (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
Nucleospin triprep kit, by Macherey-Nagel (1 mentions)
Kapa sybr fast qpcr master mix, by Roche (1 mentions)
Dharmafect 2, by Horizon Discovery (1 mentions)
Lipofectamine 2000, by Horizon Discovery (1 mentions)
Sirnas, by Horizon Discovery (1 mentions)
Normal human epidermal melanocytes, by Lonza (1 mentions)
Lenticrispr v2 vector, by Addgene (1 mentions)
Eg7 cells, by American Type Culture Collection (1 mentions)
Phage ef1al egfp w, by Addgene (1 mentions)
C57bl 6, by American Type Culture Collection (1 mentions)
C57bl 6, by Jackson ImmunoResearch (1 mentions)
Anti pd l1 10f 9g2, by BioXCell (1 mentions)
Raw264.7 cell, by American Type Culture Collection (1 mentions)
Gibco dulbecco s modified eagle s medium dmem, by Thermo Fisher Scientific (1 mentions)
Lo2 cells, by American Type Culture Collection (1 mentions)
Pen strep, by Thermo Fisher Scientific (1 mentions)
13 protocols using b16 cells
1
Culturing Murine Melanoma Cells
2
Cell Culture Protocol for B16 Melanoma and NHEM
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B16 cells were obtained from ATCC. Primary human melanocytes (Normal Human Epidermal Melanocytes, NHEM) were obtained from Lonza. All cell culture reagents were obtained from GIBCO Life technologies. siRNAs and shRNA were purchased from Dharmacon (GE Healthcare) and transfections were performed using Dharmafect II for siRNAs and Lipofectamine 2000 for shRNAs. Midiprep plasmid preparation was performed using Qiagen Midi DNA kit. KAPA SYBR FAST qPCR Master Mix was obtained from KAPA Biosystems. Genomic DNA and RNA isolations were performed using Macherey Nagel NucleoSpin® TriPrep kit (Cat no.: 740966).
3
OVA Antigen Expression in Melanoma Cells
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The OVA coding region (GenBank: V00383.1 ) was amplified from the OVA gene-transduced mouse T lymphoma cell line EG7 cells (ATCC). The cDNA was subcloned into pMXs-IRES-neo (pMXs-IN).63 (link) The gene was retrovirally transduced into mouse melanoma cell line B16 cells (ATCC). To generate β2m KO OVA/B16 cells, mouse β2m gRNA 5′-ctggtgcttgtctcactgac-3′, designed using the software program CRISPRdirect,66 (link) was subcloned into lentiCRISPRv2 vector, a gift from Feng Zhang (addgene #52961). The lentivirus was prepared and used to infect OVA/B16 cells according to our previous report.67 (link) The mutant sequence is shown in Figure S3 A. SCT encoding the leader sequence of β2m followed by SIINEKL sequence, a first linker of GGGAS (G4S)2, β2m sequence, the second liker of (G4S)4, and Kb sequence,68 (link) was subcloned into pHAGE-EF1α lentiviral vector after removing EGFP from pHAGE-EF1aL-eGFP-W (Addgene plasmid #126686), resulting in the generation of SCT/pHAGE-EF1α vectors. The lentivirus was prepared and used to infect BW5147 cells according to our previous report.67 (link) Cell lines used for the split-GFP system are described below.
4
Establishing Lung Tumor Model in Mice
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B16 cells were obtained from ATCC and were tested for their ability to form lung tumor foci upon intravenous injection into C57BL/6 mice. C57BL/6, Rag1−/− and Pmel-1 TCR transgenic mice34 (link) were purchased from the Jackson Laboratory (Bar Harbor, ME, USA). For all experiments 6- to 8-week-old mice were used. Animal experiments were carried out with approval of the Université de Sherbrooke Ethics Committee for Animal Care and Use.
5
B16 Melanoma Cell Culture
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B16 cells were purchased from ATCC. Cells were cultured in RPMI 1640 medium supplement with 10% FBS at 37 °C in 5% CO2 incubator.
6
Subcutaneous Tumor Induction and Vaccination
7
Synthesis and Characterization of Multifunctional Nanoparticles
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HAuCl4·3H2O, sodium citrate, 1-adamantanethiol, Cy5-PEG (2k)-SH, and FITC-PEG (2k)-SH were all procured from Aladdin (China). Zhiyuan Biological Technology Co. Ltd. (China) provided mono(6-mercapto-6-deoxy)-β-CD. Fetal bovine serum (FBS) and Gibco Dulbecco’s modified Eagle’s medium (DMEM) were obtained from Gibco (China). All other chemical reagents were supplied by Aladdin (China).
B16 cells, LO2 cells, and RAW264.7 cells were obtained from American Type Culture Collection (ATCC; USA). All of these cell lines were authenticated by DNA fingerprinting, isozyme detection, viability test, and mycoplasma detection. E. coli (ATCC 33694) was purchased from ATCC. Male C57BL/6 mice (6 weeks) were purchased from Faculty of Health Sciences, University of Macau. All animal procedures were approved by the Animal Ethics Committee, University of Macau and were conducted in accordance with the Animal Management Rules of the Ministry of Health of the P. R. China.
B16 cells, LO2 cells, and RAW264.7 cells were obtained from American Type Culture Collection (ATCC; USA). All of these cell lines were authenticated by DNA fingerprinting, isozyme detection, viability test, and mycoplasma detection. E. coli (ATCC 33694) was purchased from ATCC. Male C57BL/6 mice (6 weeks) were purchased from Faculty of Health Sciences, University of Macau. All animal procedures were approved by the Animal Ethics Committee, University of Macau and were conducted in accordance with the Animal Management Rules of the Ministry of Health of the P. R. China.
8
Human HNSCC Cell Line Maintenance
9
Transient Transfection of B16 and Fibroblast Cells
10
Culturing Mouse Melanoma and Monkey Kidney Cells
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Mouse melanoma B16 cells and monkey kidney Vero cells were obtained from the American Type Culture Collection (ATCC, Manassas, VA). B16 and Vero cells were maintained as a monolayer in Dulbecco’s modified Eagle’s medium (DMEM, Gibco BRL, Grand Island, NY) supplemented with 10% FBS, 100 U/ml penicillin and 100 μg/ml streptomycin (GE Healthcare, Utah, USA) at 37 °C in a humidified atmosphere of 5% carbon dioxide (CO2). The medium was refreshed every 2–3 days. B16 and Vero cells were sub-cultured using 0.25% trypsin-EDTA when the cells reached about 70% confluence.
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