| 產(chǎn)品名稱(chēng) |
HCN-2 |
| 商品貨號(hào) |
B164579 |
| Organism |
Homo sapiens, human |
| Tissue |
brain |
| Cell Type |
cortical neuron |
| Product Format |
frozen |
| Morphology |
neuronal |
| Culture Properties |
adherent |
| Biosafety Level |
1
Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country. |
| Disease |
encephalitis |
| Age |
7 years |
| Gender |
female |
| Storage Conditions |
liquid nitrogen vapor phase |
| Disclosure |
This material is cited in a US or other Patent and may not be used to
infringe the claims. Depending on the wishes of the Depositor, ATCC may be
required to inform the Patent Depositor of the party to which the material was
furnished. This material may not have been produced or characterized by ATCC. |
| Derivation |
The HCN-2 cell line was derived from cortical tissue removed from a patient undergoing hemispherectomy for intractable seizures associated with Rasmussen's encephalitis. |
| Clinical Data |
female
7 years |
| Genes Expressed |
tubulin; neurofilament protein; somatostatin; cholecystokinin-8 |
| Cellular Products |
tubulin; neurofilament protein; somatostatin; cholecystokinin-8 |
| Comments |
HCN-2 cells can be induced to differentiate when cultured with a mixture of nerve growth factor (NGF), dibutyryl cyclic adenosine monophosphate (cAMP) and 1-isobutyl-3-methylxanthine (IBMX).
Differentiation is accompanied by mature morphology and slowing of growth (doubling time greater than 120 hours). The growth rate of HCN-2 cells is stimulated by treatment with phorbol esters. A similar line (HCN-1A) is available as ATCC CRL-10442.
The cells stain positively for a number of neuronal markers including neurofilament protein, neuron specific enolase (NSE).
They are also positive for tubulin, vimentin, somatostatin (SST), glutamate, gamma aminobutyric acid (GABA), cholecystokinin - 8 (CCK-8) and vasoactive intestinal peptide (VIP).
The cells are negative for glial fibrillary acidic protein (GFAP) and myelin basis protein (MBP).
CRL-10742 has been shown to senesce at approximately passage 21. Current distribution stocks are prepared with only 4-5 Population Doublings (PDLs) remaining under recommended culture conditions after cryopreservation. |
| Complete Growth Medium |
Dulbecco's modified Eagle's medium with 4 mM L-glutamine adjusted to contain 1.5 g/L sodium bicarbonate, 4.5 g/L glucose, 90%; fetal bovine serum, 10%
|
| Subculturing |
Volumes are given for a 75 cm2 flask. Increase or decrease the amount of dissociation medium needed proportionally for culture vessels of other sizes.
NOTE: This cell line is extremely slow growing. May not be able to subculture for 12 to 14 days after recovery. Cultures do not reach 100% confluence (about 80-90%). A change in the fetal bovine serum may help increase the growth rate. Subculture at no higher than a 1:3 ratio every 10 to 12 days when cultures are about 90% confluent.
- 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 that 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.
-
To remove trypsin-EDTA solution, transfer cell suspension to centrifuge tube and spin at approximately 125 xg for 5 to 10 minutes.
Discard supernatant and resuspend cells in fresh growth medium. Add appropriate aliquots of cell suspension to new culture vessels.
- Incubate cultures at 37°C.
Subcultivation Ratio: A subcultivation ratio of 1:2 to 1:3 is recommended
Medium Renewal: Every 2 to 3 days |
| Cryopreservation |
Freeze Medium: Complete growth medium supplemented with 5% (v/v) DMSO
Storage Temperature: Liquid nitrogen vapor phase |
| Culture Conditions |
Atmosphere: Air, 95%; carbon dioxide (CO2), 5%
Temperature: 37°C |
| STR Profile |
Amelogenin: X CSF1PO: 10,15 D13S317: 11,12 D16S539: 8,11 D5S818: 13 D7S820: 9,11 THO1: 6,9.3 TPOX: 8 vWA: 14,20 |
| Name of Depositor |
Johns Hopkins University |
| U.S. Patent Number |
|
| Passage History |
CRL-10742 has been shown to senesce at approximately passage 21. Current distribution stocks are prepared with only 4-5 Population Doublings (PDLs) remaining under recommended culture conditions after cryopreservation. |
| References |
Ronnett GV, et al. Human neuronal cell line. US Patent 5,196,315 dated Mar 23 1993
|
