NeHepLxHT細(xì)胞

生長狀態(tài)： 貼壁生長

年限： neonatal

運(yùn)輸方式： 凍存運(yùn)輸

ATCC Number： CRL-4020?

細(xì)胞形態(tài)： 上皮樣

數(shù)量： 大量

細(xì)胞類型： 其他細(xì)胞類型

是否是腫瘤細(xì)胞： 0

物種來源： 人

Designations: NeHepLxHT

Depositors: ATCC

Biosafety Level: 2 [cells containing SV40 viral DNA sequences ]

Shipped: frozen

Medium & Serum: See Propagation

Growth Properties: NeHepLxHT細(xì)胞adherent

Organism: Homo sapiens

Morphology: epithelial-like

Source: Cell Type: neonatal hepatocyte

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Applications: 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.

Telomerase activity is elevated compared to the parental cells.

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.

Antigen Expression: Positive for epithelial marker pan-cytokeratin (immunocytochemistry); a subpopulation of cells is positive for albumin (flow cytometry).

NeHepLxHT細(xì)胞DNA Profile (STR): D5S818: 11, 12

D13S317: 12

D7S820: 8, 13

D16S539: 10, 11

vWA: 16, 18

THO1: 6

TPOX: 8, 11

CSF1PO: 12, 13

Amelogenin: XY

Cytogenetic Analysis: This is a diploid human cell line of male origin with a modal chromosome number of 46. However, approximately 20% of the cells have lost the Y-chromosome. Cells with marker chromosomes appeared infrequently and no consistent marker chromosomes were detected.

Age: neonatal

Gender: male

Comments: NeHepLxHT was developed from human neonatal hepatocytes by transduction with a retroviral expression vector (pLXSN) containing the hTERT gene. NeHepLxHT細(xì)胞The cell line was continuously cultured for more than twenty five passages without senescence whereas the parental cells senesced within three passages. Telomerase activity is elevated compared to the parental cells. A diploid karyotype is maintained. The gene expression profile is consistent with hepatic progenitor cells; mRNA is expressed for Alpha1-antitrypsin, Epithelial cell adhesion molecule (ECAM) and Neuronal cell adhesion molecule (NCAM).

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 is ATCC-formulated DMEM:F12 Medium Catalog No. 30-2006. To make the complete growth medium, add the following components to the base medium:

0.1 microliter/ ml Dexamethasone from HMM Hepatocyte Medium SingleQuot (Lonza/ Clonetics Catalog # CC-4192)

0.11 microliter/ ml insulin from HMM Hepatocyte Medium SingleQuot (Lonza/ Clonetics Catalog # CC-4192)

50 microgram/ ml G-418

heat-inactivated fetal bovine serum to a final concentration of 10%

Temperature: 37.0°C

Subculturing: Note: NeHepLxHT細(xì)胞The culture flasks should be pre-coated with Bovine Collagen Type I. For example, coat 0.4ml/T-25cm2 flask with 0.1% solution (Sigma Cat # C8919) at 4?C for at least 18 hours. Before introducing cells, remove excess solution and rinse flask 3 times with a balanced salt solution.Protocol: Volumes used in this protocol are for 75 sq cm flasks; 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 0.05%Trypsin-0.53mMEDTA 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 37C to facilitate dispersal.

Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.

Transfer cell suspension to a centrifuge tube and spin at approximately 125 X g for 5 to 10 minutes. Discard supernatant.

Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new coated culture vessels. An inoculum of 4 X 10(3) to 6 X 10(3) viable cells/sq. cm is recommended.

Incubate cultures at 37C. Subculture when cell concentration is between 2 X 10(4) and 6X 10(4) cells/sq. cm.

Subcultivation Ratio: 1:5 to 1:10

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

Storage temperature: liquid nitrogen vapor phase

Doubling Time: NeHepLxHT細(xì)胞about 65 hours