產(chǎn)品名稱 |
hTERT-HME1 [ME16C] |
商品貨號 |
B212032 |
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]
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 |
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. |
Images |
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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.
- Remove and discard culture medium.
- 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.
- To remove trypsin-EDTA solution, add 2.0 to 3.0 mL of 0.1% SoybeanTrypsin Inhibitor solution and aspirate cells by gently pipetting.
- Transfer cell suspension to a 15 mL centrifuge tube and spin at approximately 125 x g for 5 to 10 minutes.
- 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.
- 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
Caputo JL. Biosafety procedures in cell culture. J. Tissue Culture Methods 11:223-227, 1988
Freshney RI. Culture of Animal Cells: A Manual of Basic Technique, 4th edition. New York: Wiley Liss; 2000. For more information on enzymatic dissociation and subculturing of cell lines see Chapter 10.
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