hTERT-HME1 [ME16C] (ATCC® CRL-4010)

Organism: Homo sapiens, human  /  Cell Type: Epithelial cells immortalized with hTERT  /  Tissue: Breast; mammary gland; Epithelium  /  Disease: Normal

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Organism Homo sapiens, human
Tissue Breast; mammary gland; Epithelium
Cell Type Epithelial cells immortalized with hTERT
Product Format frozen
Morphology Epithelial-like
Culture Properties Adherent
Biosafety Level 2 [cells contain SV40 viral DNA sequences]
Disease Normal
Age 53 years
Gender Female
Storage Conditions Liquid nitrogen vapor phase
Karyotype This is a pseudo-diploid cell line of female origin with a modal chromosome count of 46 and a low-to-moderate rate of polyploidy. However, even though the line generally has 46 chromosomes per cell, several of those 46 were derivative or marker chromosomes. There were two copies of a karyotypically normal X-chromosome present in 50-60% of the cells. Other features included a normal variation in the heterochromatic region of chromosome 1 (1qh+), a consistent derivative-10 marker chromosome (present in most cells) and 2 other markers: del(3)(p24?) and del(16)(q21~23?) (present in approximately 20-30% of the analyzed cells). Overall, approximately 3-8 marker chromosomes were present in the analyzed metaphase spreads and satellite associations appeared sporadically.
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Derivation

Human mammary epithelium, HME1 cell line, was derived from a 53-year-old patient undergoing reduction mammoplasty surgery (no history of breast cancer). 

The HME1 cells were immortalized by infection with the retrovirus pBabepuro+hTERT vector and cultured in complete growth medium containing puromycin until stable clones were selected. 

Antigen Expression

Positive for the cytokeratin epithelial marker (immunocytochemistry) and for the Mucin 1, transmembrane (MUC1) protein (Homo sapiens; detection by flow cytometry). 

Complete Growth Medium The base medium for this cell line (MEBM) along with all the additives can be obtained from Lonza/Clonetics Corporation as a kit: MEGM, Kit Catalog No. CC-3150. ATCC does not use the GA-1000 (gentamycin-amphotericin B mix) provided with kit. Note: Do not filter complete medium
Subculturing Volumes are given for a 75 cm2 flask; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.
  1. Remove and discard culture medium.
  2. Add 2.0 to 3.0 mL of 0.25% (w/v) Trypsin - 0.53 mM EDTA solution to the flask and observe cells under an inverted microscope until the 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.
  3. To remove trypsin-EDTA solution, add 2.0 to 3.0 mL of 0.1% SoybeanTrypsin Inhibitor solution and aspirate cells by gently pipetting.
  4. Transfer cell suspension to a 15 mL centrifuge tube and spin at approximately 125 x g for 5 to 10 minutes.
  5. Discard supernatant and resuspend cells in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 7 x 103 to 9x 103 viable cells/cm2  is recommended. Subculture cultures when they reach a cell concentration between 4 x 104 and 6 x 104 cells/cm2.
  6. Incubate cultures at 37°C.
Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:6 is recommended.
Medium Renewal: Every 2 to 3 days
Note:For more information on enzymatic dissociation and subculturing of cell lines consult Chapter 12 in Culture of Animal Cells: A Manual of Basic Technique by R. Ian Freshney.
Cryopreservation
Culture medium, 90%; DMSO, 10%

Culture Conditions

Temperature: 37°C
Atmosphere: Air, 95%; Carbon dioxide (CO2), 5%


STR Profile
Amelogenin: X
CSF1PO: 10
D13S317: 11,12
D16S539: 11,12
D5S818: 11
D7S820: 7,12
THO1: 7,8
TPOX: 10,12
vWA: 15,16
Population Doubling Level (PDL) As part of our quality control, we have tested this cell line for its ability to grow for a minimum of 15 population doublings after recovery from cryopreservation. We have also compared its karyotype, telomerase expression level, growth rate, morphology and tissue-specific markers when first recovered from cryopreservation with that of cells at 10+ population doublings to ensure that there is no change in these parameters and that the cells are capable of extended proliferation.
Name of Depositor JW Shay
References

Yi X, et al. Both transcriptional and posttranscriptional mechanisms regulate human telomerase template RNA levels. Mol. Cell. Biol. 19(6): 3989-3997, 1999. PubMed: 10330139

Morgenstern JP, Land H. Advanced mammalian gene transfer: high titre retroviral vectors with multiple drug selection markers and a complementary helper-free packaging cell line. Nucleic Acids Res. 18: 3587-3596, 1990. PubMed: 2194165

Shay JW, et al. E6 of human papillomavirus type 16 can overcome the M1 stage of immortalization in human mammary epithelial cells but not in human fibroblasts. Oncogene 8: 1407-1413, 1993. PubMed: 8389027

Gollahon LS, Shay JW. Immortalization of human mammary epithelial cells transfected with mutant p53 (273his). Oncogene 12: 715-725, 1996. PubMed: 8632893

Van Der Haegen BA, Shay JW. Immortalization of human mammary epithelial cells by SV40 large T-antigen involves a two step mechanism. In Vitro Cell. Dev. Biol. 29: 180-182, 1993. PubMed: 8463179

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.
  • License agreement required for commercial customer uses.
  • This material is distributed for research purposes only. A signed addendum to the ATCC Material Transfer Agreement must be sent to ATCC in advance of shipment.
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This material requires that the Addendum for Commercial and For-Profit Organizations or the Addendum for Noncommercial and Academic Organizations be signed and returned to ATCC before shipment. The price listed above is for noncommercial and academic organizations only. Commercial and for-profit organizations should call for pricing.  

References

Yi X, et al. Both transcriptional and posttranscriptional mechanisms regulate human telomerase template RNA levels. Mol. Cell. Biol. 19(6): 3989-3997, 1999. PubMed: 10330139

Morgenstern JP, Land H. Advanced mammalian gene transfer: high titre retroviral vectors with multiple drug selection markers and a complementary helper-free packaging cell line. Nucleic Acids Res. 18: 3587-3596, 1990. PubMed: 2194165

Shay JW, et al. E6 of human papillomavirus type 16 can overcome the M1 stage of immortalization in human mammary epithelial cells but not in human fibroblasts. Oncogene 8: 1407-1413, 1993. PubMed: 8389027

Gollahon LS, Shay JW. Immortalization of human mammary epithelial cells transfected with mutant p53 (273his). Oncogene 12: 715-725, 1996. PubMed: 8632893

Van Der Haegen BA, Shay JW. Immortalization of human mammary epithelial cells by SV40 large T-antigen involves a two step mechanism. In Vitro Cell. Dev. Biol. 29: 180-182, 1993. PubMed: 8463179