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CCD 1106 KERTr (ATCC® CRL-2309™)

Organism: Homo sapiens, human  /  Cell Type: keratinocyte; human papillomavirus 16 (HPV-16) E6/E7 transformed  /  Tissue: skin  / 

Permits and Restrictions

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Organism Homo sapiens, human
Tissue skin
Cell Type keratinocyte; human papillomavirus 16 (HPV-16) E6/E7 transformed
Product Format frozen
Morphology epithelial
Biosafety Level 2
[Cells may contain the Human Papilloma viral (HPV) sequences]

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.
Age 133 days gestation
Storage Conditions liquid nitrogen vapor phase
Derivation Passage 2 cells were transformed with a retrovirus vector (LXSN16E6E7 produced by ATCC CRL-2203) in the presence of polybrene.
Antigen Expression
epithelial specific antigen
Oncogene E6/E7 +
Genes Expressed
E6/E7 +, epithelial specific antigen
Comments

This line stains positively with antibody for cytokeratin.

After 50 population doublings, the cells continue dividing and retain cuboidal morphology.

E6E7 sequences were detected by PCR in cells at passage 18.

Major Histocompatibility Complex class I or II molecules were not expressed on these cells, but PCR analyses revealed presence of the genes for directing synthesis of HLA antigens.

Epithelial specific antigen was detected using antibody produced by the Ep16 hybridoma (ATCC HB-155).
Complete Growth Medium These cells are grown in Keratinocyte-Serum Free Medium (Gibco 17005-042) with added Keratinocytes Supplements (Gibco 37000-015) including Bovine Pituitary Extract (BPE; Gibco 13028-014) and human recombinant epidermal growth factor (EGF; Gibco 10450-013) further supplemented with:
  • Additional 30 ng/ml human recombinant epidermal growth factor (EGF; BD cat# 354052)

Do not filter the complete medium.

This medium is formulated for use with a 5% CO2 in air atmosphere.
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.

  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. To remove trypsin-EDTA solution, transfer cell suspension to centrifuge tube and spin at approximately 125 x g for 5 to 10 minutes.
  6. Discard supernatant and resuspend cells in fresh growth medium.  Add appropriate aliquots of cell suspension to new culture vessels.
  7. Place culture vessels in incubators at 37°C.

Subculture Ratio: 1:3 to 1:5
Medium Renewal: Twice a week.
Note: For more information on enzymatic dissociation and subculturing of cell lines consult Chapter 10 in Culture of Animal Cells, a manual of Basic Technique by R. Ian Freshney, 3rd edition, published by Alan R. Liss, N.Y., 1994.
Cryopreservation
Freeze Medium: Ham's F12 medium 85%; DMSO, 5%; FBS 10%
Storage Temperature: Liquid nitrogen vapor phase
Culture Conditions
Temperature: 37°C
Add fresh medium twice per week
Name of Depositor L Vilner
Passage History
E6E7 sequences were detected by PCR in cells at passage 18.
Passage 2 cells were transformed with a retrovirus vector ( LXSN16E6E7 produced by ATCC CRL-2203) in the presence of polybrene.
References

Zabrenetzky V, et al. The isolation, immortalization and characterization of human fetal keratinocytes. In Vitro Cell. Dev. Biol. 33: Part II, p. 34A, 1997.

Basic Documentation
Product Sheet
Product Sheet
Certificate of Analysis
Certificate of Analysis
SDS
SDS
Restrictions

Note: These cells are distributed for research purposes only. The American Type Culture Collection ( ATCC ) releases the line subject to the following: 1. CCD 1106 KERTr cells or their products must not be distributed to third parties. Commercial interests are the exclusive property of the ATCC; 2. Any proposed commercial use of this cell line must first be negotiated with the ATCC, P.O. Box 1549, Manassas VA 20108, telephone 703-365-2704, FAX 703-365-2730.
References

Zabrenetzky V, et al. The isolation, immortalization and characterization of human fetal keratinocytes. In Vitro Cell. Dev. Biol. 33: Part II, p. 34A, 1997.