EML4-ALK Fusion-A549 Isogenic-Luc2 (ATCC® CCL-185IG-LUC2)

Organism: Homo sapiens, human  /  Tissue: lung  /  Disease: carcinoma

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Organism Homo sapiens, human
Tissue lung
Product Format frozen 1.0 mL
Morphology epithelial-like
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.

Disease carcinoma
Age 58 years
Gender male
Ethnicity Caucasian
Applications EML4-ALK positive lung cancer model. Excellent signal/background ratio and stable luciferase expression make this cell line ideal for in vivo bioluminescence imaging of xenograft animal models to study human lung cancer and monitor the activity of anti-cancer drugs. It also can be used in cell-based assays for cancer research.
Storage Conditions liquid nitrogen vapor phase
Tumorigenic Yes, tested in NSG mice
Comments

This luciferase-expressing cell line was derived from parental line CCL-185IG by transduction with lentiviral vector encoding firefly luciferase gene (luc2) under control of EF-1 alpha promoter. The parental line CCL-185IG (ATCC CCL-185IG) is a CRISPR gene-edited non-small cell lung cancer model containing EML4-ALK gene fusion. 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 (8 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.

Complete Growth Medium The base medium for this cell line is F-12K Medium (ATCC 30-2004). 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 8 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 103 and 1 x 104 viable cells/cm2. Do not exceed 7 x 104 cells/cm2.
  6. Incubate cultures at 37°C.
Interval: Maintain cultures at a cell concentration between 6 X 103 and 6 X 104 cell/cm2.
Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:5 is recommended
Medium Renewal: 2 to 3 times per week
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 Approximately 1 x 106 cells
Volume 1.0 mL
STR Profile
Amelogenin: X
CSF1PO: 10,12
D13S317: 11
D16S539: 11,12
D5S818: 11
D7S820: 8,11
THO1: 8,9.3
TPOX: 8,11
vWA: 14
Sterility Tests Bacteria and yeast: No growth
Mycoplasma: No growth
Viral Testing Hepatitis B: None detected
Cytomegalovirus: None detected
Human immunodeficiency virus: None detected
Epstein-Barr virus: None detected
Human papillomavirus: None detected
Functional Tests Luciferase activity: signal to noise ≥ 1,000
In Vitro Luminesence: ≥ 500,000 photons/cell/sec, subject to imaging and culturing conditions. Genotype testing for gene fusion: EML4-ALK fusion product detected by PCR.
Population Doubling Time Approximately 23 hours
Name of Depositor ATCC
Year of Origin 2017
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

Giard DJ, et al. In vitro cultivation of human tumors: establishment of cell lines derived from a series of solid tumors. J. Natl. Cancer Inst. 51: 1417-1423, 1973. PubMed: 4357758

Gershwin ME, et al. Immunobiology of heterotransplanted human tumors in nude mice. J. Natl. Cancer Inst. 58: 1455-1463, 1977. PubMed: 857033

Cormier JN, et al. Heterogeneous expression of melanoma-associated antigens and HLA-A2 in metastatic melanoma in vivo. Int. J. Cancer 75: 517-524, 1998. PubMed: 9466650

Sharma SD, et al. Melanotropic peptide-conjugated beads for microscopic visualization and characterization of melanoma melanotropin receptors. Proc. Natl. Acad. Sci. USA 93: 13715-13720, 1996. PubMed: 8943000

Basic Documentation
Other Documentation
Restrictions

This material is subject to the following restrictions in addition to those outlined in the ATCC Material Transfer Agreement: Luciferase Label License.
For information on obtaining additional rights, please contact:
ATCC Licensing
Email: licensing@atcc.org

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.

In addition to the foregoing, this product's use is governed by the CRISPR Label License Agreement. For information on purchasing a license to use this product for purposes other than those permitted in the CRISPR Label License Agreement, please contact The Broad Institute at partnering@broadinstitute.org.

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

Giard DJ, et al. In vitro cultivation of human tumors: establishment of cell lines derived from a series of solid tumors. J. Natl. Cancer Inst. 51: 1417-1423, 1973. PubMed: 4357758

Gershwin ME, et al. Immunobiology of heterotransplanted human tumors in nude mice. J. Natl. Cancer Inst. 58: 1455-1463, 1977. PubMed: 857033

Cormier JN, et al. Heterogeneous expression of melanoma-associated antigens and HLA-A2 in metastatic melanoma in vivo. Int. J. Cancer 75: 517-524, 1998. PubMed: 9466650

Sharma SD, et al. Melanotropic peptide-conjugated beads for microscopic visualization and characterization of melanoma melanotropin receptors. Proc. Natl. Acad. Sci. USA 93: 13715-13720, 1996. PubMed: 8943000