HCN-2 (ATCC® CRL-10742)

Organism: Homo sapiens, human  /  Cell Type: cortical neuron  /  Tissue: brain  /  Disease: encephalitis

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Organism Homo sapiens, human
Tissue brain
Cell Type cortical neuron
Product Format frozen
Morphology neuronal
Culture Properties adherent
Biosafety Level 1
Disease encephalitis
Age 7 years
Gender female
Storage Conditions liquid nitrogen vapor phase
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.

  1. 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.
  2. 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.
  3. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
  4. 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.
  5. 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
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

Notice: Necessary PermitsPermits

These permits may be required for shipping this product:

  • Customers located in the state of Hawaii will need to contact the Hawaii Department of Agriculture to determine if an Import Permit is required. A copy of the permit or documentation that a permit is not required must be sent to ATCC in advance of shipment.
Basic Documentation
References

Ronnett GV, et al. Human neuronal cell line. US Patent 5,196,315 dated Mar 23 1993