B16-F10-Luc2 (ATCC® CRL-6475-LUC2)

Organism: Mus musculus, mouse  /  Tissue: skin  / 

Permits and Restrictions

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Organism Mus musculus, mouse
Tissue skin
Product Format frozen 1.0 mL
Morphology mixture of spindle-shaped and epithelial-like cells
Culture Properties adherent
Biosafety Level 2

Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.

Strain C57BL/6J
Applications Excellent signal/background ratio and stable Luciferase expression make this cell line ideal for in vivo bioluminescence imaging of xenograft animal model to study human cancer and monitor activity of anti-cancer drug. It also can be used in cell-based assays for cancer research.
Storage Conditions liquid nitrogen vapor phase
Tumorigenic Yes, tested in C57/BL6
Comments

This luciferase expressing cell line was derived from parental line CRL-6323 by transduction with lentiviral vector encoding firefly luciferase gene (luc2) under control of EF-1 alpha promoter. This cell line was established through single cell cloning, and the cells constitutively express high levels of enzymatically active luciferase protein, which can be detected via in vitro and in vivo bioluminescence assays. The cells should be maintained in Blasticidin (10 µg/mL) containing medium in routine cell culture. It is recommended to remove Blasticidin prior to and during the experiment procedure when the cells are injected into animals in vivo, or co-cultured with other cell types in vitro.

Note: This cell line produces melanin. This may cause the culture medium or cells to appear dark brown or black, especially when the cultures are approaching a high level of confluence.

Complete Growth Medium

The base medium for this cell line is Dulbecco's Modified Eagle's Medium, (DMEM; ATCC 30-2002). To make the complete growth medium, add the following components to the base medium:

  • Fetal bovine serum (FBS; ATCC 30-2020) to a final concentration of 10%
  • Blasticidin to a final concentration of 10 µg/mL
Subculturing
Volumes used in this protocol are for 75 cm2 flask; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes. Corning® T-75 flasks (catalog #430641) are recommended for subculturing this product.
  1. Remove and discard culture medium.
  2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin- 0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.
  3. Add 2.0 to 3.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).
    Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.
  4. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
  5. Add appropriate aliquots of the cell suspension to new culture vessels.
    Cultures can be established between 2 x 104 and 4 x 104 viable cells/cm2.
  6. Incubate cultures at 37°C.
div>Interval: Maintain cultures at a cell concentration between 1 X 104 and 2.3 X 105 cell/cm2.
Subcultivation Ratio: A subcultivation ratio of 1:10 is recommended
Medium Renewal: Every2 to 3 days
Cryopreservation Complete growth medium supplemented with 5% (v/v) DMSO (ATCC 4-X)
Culture Conditions
Atmosphere: air, 95%; carbon dioxide (CO2), 5%
Temperature: 37°C
Cells per Vial ≥ 1.0 x 106
Volume 1.0 mL
Sterility Tests Bacteria and yeast: No growth
Mycoplasma: No growth
Functional Tests Luciferase activity: signal to noise ≥ 1,000 RLUs
In Vitro Luminesence: 20,000 photons/cell/sec, subject to imaging and culturing conditions
Population Doubling Time approximately 12 hrs
Name of Depositor ATCC
Year of Origin 2018
References

Zinn KR, et al. Noninvasive bioluminescence imaging in small animals. ILARJ 49: 103-115, 2008. PubMed: 18172337

Dothager RS, et al. Advances in bioluminescence imaging of live animal models. Curr Opin Biotechnol 20: 45-53, 2009. PubMed: 19233638

Fidler IJ. Biological behavior of malignant melanoma cells correlated to their survival in vivo. Cancer Res. 35: 218-224, 1975. PubMed: 1109790

Fidler IJ, et al. Tumoricidal properties of mouse macrophages activated with mediators from rat lymphocytes stimulated with concanavalin A. Cancer Res. 36: 3608-3615, 1976. PubMed: 953987

Fidler IJ, Bucana C. Mechanism of tumor cell resistance to lysis by syngeneic lymphocytes. Cancer Res. 37: 3945-3956, 1977. PubMed: 908034

Fidler IJ, Kripke ML. Metastasis results from preexisting variant cells within a malignant tumor. Science 197: 893-895, 1977. PubMed: 887927

Briles EB, Kornfeld S. Isolation and metastatic properties of detachment variants of B16 melanoma cells. J. Natl. Cancer Inst. 60: 1217-1222, 1978. PubMed: 418183

Fidler IJ. Selection of successive tumour lines for metastasis. Nat. New Biol. 242: 148-149, 1973. PubMed: 4512654

Li M, et al. Loss of intracisternal A-type retroviral particles in BL6 melanoma cells transfected with MHC class I genes. J.Gen. Virol. 77: 2757-2765, 1996. PubMed: 8922469

Notice: Necessary PermitsPermits

These permits may be required for shipping this product:

  • Customers located in the state of Hawaii will need to contact the Hawaii Department of Agriculture to determine if an Import Permit is required. A copy of the permit or documentation that a permit is not required must be sent to ATCC in advance of shipment.
Basic Documentation
Other Documentation
Restrictions

This material’s use is governed by the NanoLuc Limited Use Label License. For information on purchasing a license to use this product for purposes other than those permitted in the Label License, please contact Promega.

For commercial accounts, this cell line is only distributed under the terms of a fully signed and executed ATCC® Material Transfer Agreement and Addendum. If the commercial account is screening per completed Addendum, the recipient will be required to pay a Screening Fee (ATCC® ACS-2103F™).

Screening Use is defined as use of Biological Material in small molecule and biologic drug discovery, including initial target identification and validation, assay development, high throughput screening, hit identification, lead optimization, and selection of candidates for clinical development.

If the commercial account is not screening per the completed Addendum, the recipient will not be required to pay a Screening Fee.

References

Zinn KR, et al. Noninvasive bioluminescence imaging in small animals. ILARJ 49: 103-115, 2008. PubMed: 18172337

Dothager RS, et al. Advances in bioluminescence imaging of live animal models. Curr Opin Biotechnol 20: 45-53, 2009. PubMed: 19233638

Fidler IJ. Biological behavior of malignant melanoma cells correlated to their survival in vivo. Cancer Res. 35: 218-224, 1975. PubMed: 1109790

Fidler IJ, et al. Tumoricidal properties of mouse macrophages activated with mediators from rat lymphocytes stimulated with concanavalin A. Cancer Res. 36: 3608-3615, 1976. PubMed: 953987

Fidler IJ, Bucana C. Mechanism of tumor cell resistance to lysis by syngeneic lymphocytes. Cancer Res. 37: 3945-3956, 1977. PubMed: 908034

Fidler IJ, Kripke ML. Metastasis results from preexisting variant cells within a malignant tumor. Science 197: 893-895, 1977. PubMed: 887927

Briles EB, Kornfeld S. Isolation and metastatic properties of detachment variants of B16 melanoma cells. J. Natl. Cancer Inst. 60: 1217-1222, 1978. PubMed: 418183

Fidler IJ. Selection of successive tumour lines for metastasis. Nat. New Biol. 242: 148-149, 1973. PubMed: 4512654

Li M, et al. Loss of intracisternal A-type retroviral particles in BL6 melanoma cells transfected with MHC class I genes. J.Gen. Virol. 77: 2757-2765, 1996. PubMed: 8922469