NE-GFP-4C cell line小鼠神经干细胞株 BioVector NTCC质粒载体菌种细胞基因保藏中心

NE-GFP-4C cell line小鼠神经干细胞株

NE-GFP-4C
BioVector - CRL-2926

Product category
Animal cells
Organism
Mus musculus, mouse
Classification
Eukaryota, Animalia, Metazoa, Chordata, Vertebrata, Tetrapod
Cell type
neural stem cell
Morphology
neuroepithelial
Tissue
Brain; Neuroectodermal
Applications
3D cell culture
Neuroscience

Growth properties
Adherent
Derivation
The neuroepithelial cell lines, NE-4C (CRL-2925) and NE-GFP-4C (CRL-2926) were established from the cerebral vesicles of 9-day-old mouse embryos lacking the functional p53 genes.
Age
embryo
Strain
C57Bl/Sv129 (p53-/-)
Antigen expression
Sox-2, Otx-2, En-1
Comments
Both NE-4C (CRL-2925) and NE-GFP-4C (CRL-2926) differentiate into 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 is capable of developing morphologically differentiated neurons. Ref Ref
Handling information
Unpacking and storage instructions
Check all containers for leakage or breakage.
Remove the frozen cells from the dry ice packaging and immediately place the cells at a temperature below _x001f_-130°C, preferably in liquid nitrogen vapor, until ready for use.
Complete 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%
Temperature
37°C
Atmosphere
95% Air, 5% CO2
Handling procedure
To insure the highest level of viability, thaw the vial and initiate the culture as soon as possible upon receipt. If upon arrival, continued storage of the frozen culture is necessary, it should be stored in liquid nitrogen vapor phase and not at -70°C. Storage at -70°C will result in loss of viability.

Note: The culture flasks should be pre-coated with 15µg/mL poly-L-lysine (Sigma Cat. #P-9155 or equivalent) at least 2 hours in advance.


Thaw the vial by gentle agitation in a 37°C water bath. To reduce the possibility of contamination, keep the O-ring and cap out of the water. Thawing should be rapid (approximately 2 minutes).
Remove the vial from the water bath as soon as the contents are thawed, and decontaminate by dipping in or spraying with 70% ethanol. All of the operations from this point on should be carried out under strict aseptic conditions.
Transfer the vial contents to a centrifuge tube containing 9.0 mL complete culture medium and spin at approximately 125 x g for 5 to 10 minutes.
Resuspend cell pellet with the recommended complete medium (see the specific batch information for the culture recommended dilution ratio) and dispense into a 25 cm2 or a 75 cm2 poly-L-lysine coated culture flask. It is important to avoid excessive alkalinity of the medium during recovery of the cells. It is suggested that, prior to the addition of the vial contents, the culture vessel containing the complete growth medium be placed into the incubator for at least 15 minutes to allow the medium to reach its normal pH (7.0 to 7.6).
Incubate the culture at 37°C in a suitable incubator. A 5% CO2 in air atmosphere is recommended if using the medium described on this product.
Subculturing procedure
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 with15 µg/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.
Reagents for cryopreservation
Complete growth medium supplemented with 10% (v/v) DMSO

Mycoplasma contamination
Not detected
Population doubling time
Approximately 16 hrs

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