hTERT-HME1 [ME16C]細(xì)胞

細(xì)胞形態(tài)： 上皮樣

細(xì)胞類型： 其他細(xì)胞類型

年限： 53 years *****

是否是腫瘤細(xì)胞： 0

物種來源： 人

ATCC Number： CRL-4010?

數(shù)量： 大量

生長(zhǎng)狀態(tài)： 貼壁生長(zhǎng)

運(yùn)輸方式： 凍存運(yùn)輸

器官來源： **

Designations: hTERT-HME1 [ME16C]

Depositors: JW Shay

hTERT-HME1 [ME16C]細(xì)胞Biosafety Level: 2 [cells containing SV40 viral DNA sequences ]

Shipped: frozen

Medium & Serum: See Propagation

Growth Properties: adherent

Organism: Homo sapiens

Morphology: epithelial

Source: Organ: mammary gland; breast

Cell Type: epithelial immortalized with hTERT

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Applications: s 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.

hTERT-HME1 [ME16C]細(xì)胞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.

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

Reverse Transcript: N

Antigen Expression: cytokeratin; Homo sapiens, expressed (cytokeratins were assayed by immunocytochemistry using a pan-cytokeratin antibody) (cytokeratins were assayed by immunocytochemistry using a pan-cytokeratin antibody)

mucin 1, transmembrane (MUC1); Homo sapiens, expressed (detection by flow cytometry) (detection by flow cytometry)

DNA Profile (STR): Amelogenin: X

CSF1PO: 10

D13S317: 11,12

D16S539: 11,12

D5S818: 11

D7S820: 7,12

THO1: 7,8

TPOX: 10,12

vWA: 15,16

Cytogenetic Analysis: hTERT-HME1 [ME16C]細(xì)胞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.

Age: 53 years *****

Gender: female

HeLa Markers: N

Comments: 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. These cells do not undergo growth arrest; in cell culture, due to the exogenous expression of the telomerase gene (1-5).

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.

Propagation: ATCC 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: hTERT-HME1 [ME16C]細(xì)胞Do not filter complete medium

Atmosphere: air, 95%; carbon dioxide (CO2), 5%

Temperature: 37.0°C

Subculturing: Protocol:

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 37C 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 10(3) to 9 X 10(3) viable cells/sq. cm is recommended. Subculture cultures when they reach a cell concentration between 4 X 10(4) and 6 X 10(4) cells/sq. cm.

Incubate cultures at 37C.

Interval: weekly

Subcultivation Ratio: hTERT-HME1 [ME16C]細(xì)胞A subcultivation ratio of 1:4 to 1:6 is recommended.

Medium Renewal: Every 2 to 3 days

Preservation: Freeze medium: culture medium, 90%; DMSO, 10%

Storage temperature: liquid nitrogen vapor phase

Doubling Time: approximately 25 hr

Related Products: 0.25% (w/v) Trypsin - 0.53 mM EDTA in Hank' BSS (w/o Ca++, Mg++):ATCC 30-2101

Cell culture tested DMSO:ATCC 4-X

Erythrosin B vital stain solution:ATCC 30-2404

plasmid in bacteria:ATCC MBA-141

References: 44175: 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

91777: 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

91857: 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

91858: Gollahon LS, Shay JW. Immortalization of human mammary epithelial cells transfected with mutant p53 (273his). Oncogene 12: 715-725, 1996. PubMed: 8632893

91859: 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