Organ: liver Cell Type: immortalized with SV40 large T antigen
Permits/Forms:
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Age:
fetus
Comments:
The cell line was derived from human fetal liver tissue, and immortalized by transformation with an ecotropic retrovirus containing a temperature sensitive SV40 T-antigen (tsA58) and a G418 resistance gene. After several days at 37C, the temperature of the medium was lowered to 32C. Cells were selected with G418. The selected cells were expanded and maintained. The cells produce flk-1 and some flk-2 ligands.
ATCC complete growth medium: Iscove's modified Dulbecco's medium with 4 mM L-glutamine adjusted to contain 1.5 g/L sodium bicarbonate supplemented with 0.1 mM non-essential amino acids, 1.0 mM sodium pyruvate, and 15% fetal bovine serum Atmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 32.0°C
Subculturing:
Protocol:
Remove and discard culture medium.
Briefly rinse the cell layer with 0.25% (w/v) Trypsin- 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor.
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 32°C to facilitate dispersal.
Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.
Add appropriate aliquots of the cell suspension to new culture vessels.
Incubate cultures at 32°C.
Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:4 is recommended Medium Renewal: Every 2 to 3 days
Preservation:
Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO Storage temperature: liquid nitrogen vapor phase
Doubling Time:
48 hours
Related Products:
Recommended medium (without the additional supplements or serum described under ATCC Medium):ATCC 30-2005
recommended serum:ATCC 30-2020
References:
70742: Lemischka IR. Nucleic acids encoding fragments of hematopoietic stem cell receptor flk-2. US Patent 5,270,458 dated Dec 14 1993 70744: Lemischka IR. Tyrosine kinase receptor flk-2 and fragments thereof. US Patent 5,367,057 dated Nov 22 1994 70745: Lemischka IR. Tyrosine kinase receptor human flk-2-specific antibodies. US Patent 5,548,065 dated Aug 20 1996 70746: Lemischka IR. Nucleic acids encoding soluble human FLK-2 extracellular domain. US Patent 5,621,090 dated Apr 15 1997 70747: Lemischka IR. Antibodies against tyrosine kinase receptor flk-1. US Patent 5,747,651 dated May 5 1998 70748: Lemischka IR. Method for isolating stem cells expressing flk-1 receptors. US Patent 5,912,133 dated Jun 15 1999
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[VIP第6年] 指数:6
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