NE-4C cell line鼠胚胎神经上皮细胞株细胞系 BioVector NTCC质粒载体菌种细胞基因保藏中心

NE-4C cell line鼠胚胎神经上皮细胞株细胞系

BioVector NTCC质粒载体菌种细胞基因保藏中心

Organism: Mus musculus, mouse
Tissue: brain, neuroectodermal
Cell Type: neural stem cell
Age: embryo
Morphology: neuroepithelial
Growth Properties: adherent
Derivation
The neuroepithelial cell lines, NE-4C (ATCC CRL-2925) and NE-GFP-4C (ATCC CRL-2926) were established from the cerebral vesicles of 9-day-old mouse embryos lacking the functional p53 genes. ­
Antigen Expression
Sox-2, Otx-2, En-1
Comments
The cells from both cell lines differentiated to neurons and astrocytes when exposed to retinoic acid. The GFP-transfected clone, NE-GFP-4C, when implanted into the forebrain of adult, new-born, and fetal mice or into the mid- and forebrain vesicles of early chick embryos was capable of developing morphologically differentiated neurons
Complete Growth Medium The base medium for this cell line is ATCC-formulated Eagle's Minimum Essential Medium, Catalog No. 30-2003. To make the complete growth medium, add the following components to the base medium:
additional 2mM L-Glutamine
fetal bovine serum (FBS) to a final concentration of 10%

Subculturing
Volumes used in this protocol are for 75 cm2 flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.
Note: The culture flasks should be pre-coated with15µg/mL poly-L-lysine (Sigma Cat #P-9155) at least 2 hours in advance.

Remove and discard culture medium.
Briefly rinse the cell layer with Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS) or 0.05% (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 37°C 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 poly-L-lysine coated culture vessels. An inoculum of 2 X 104 to 4 X 104 viable cells/cm2 is recommended.
Incubate cultures at 37°C. Subculture when cell concentration is between 3 X 105 and 4 X 105 cells/cm2.

Subcultivation ratio: A subcultivation ratio of 1:4 to 1:10 is recommended.
Medium renewal: Every 2 to 3 days.

Cryopreservation
Freeze medium: Culture medium, 90%; DMSO,10%
Storage temperature: liquid nitrogen vapor phase
Culture Conditions
Atmosphere: 5% CO2 in air recommended
Temperature: 37°C
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