The base medium for this cell line is ATCC-formulated DMEM:F12 Medium Catalog No. 30-2006.To make the complete growth medium, add the following components to the base medium:
- extra 10 mM HEPES (for a final conc. of 25 mM)
- 10 ng/ml cholera toxin
- 0.005 mg/ml insulin
- 0.005 mg/ml transferrin
- 100 ng/ml hydrocortisone
- fetal bovine serum 10%(final conc.
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| Cryopreservation |
Volumes are given for a 75 cm2 flask. Increase or decrease the amount of dissociation medium needed proportionally for culture vessels of other sizes.
- 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. An inoculum of 4 to 6 X 103 viable cell/cm2 is recommended.
- Incubate cultures at 37°C.
Subcultivation Ratio: A subcultivation ratio of 1:2 to 1:4 is recommended every 10 to 15 days
Medium Renewal: Every 3 to 4 days. |
| Name of Depositor |
Amelogenin: X,Y CSF1PO: 11,12 D13S317: 12,13 D16S539: 9,11 D5S818: 12,13 D7S820: 8,12 THO1: 6,9.3 TPOX: 8,11 vWA: 16 |
