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Propagation:
ATCC complete growth medium: Dulbecco's modified Eagle's medium with 4 mM L-glutamine adjusted to contain 1.5 g/L sodium bicarbonate and 4.5 g/L glucose and supplemented with 0.1 Unit/ml bovine insulin, 95%; fetal bovine serum, 5% Atmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37.0°C
Subculturing:
Protocol:
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 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 37C to facilitate dispersal.
Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.
Add appropriate aliquots of the cell suspension to new culture vessels.
Incubate cultures at 37C.
Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:6 is recommended Medium Renewal: Twice per week
Recommended medium (without the additional supplements or serum described under ATCC Medium):ATCC 30-2002
recommended serum:ATCC 30-2020
0.25% (w/v) Trypsin - 0.53 mM EDTA in Hank' BSS (w/o Ca++, Mg++):ATCC 30-2101
Cell culture tested DMSO:ATCC 4-X
References:
22491: Quaroni A, et al. Epithelioid cell cultures from rat small intestine. Characterization by morphologic and immunologic criteria. J. Cell Biol. 80: 248-265, 1979. PubMed: 88453 26096: Quaroni A, Isselbacher KJ. Cytotoxic effects and metabolism of benzo[a]pyrene and 7,12- dimethylbenz[a]anthracene in duodenal and ileal epithelial cell cultures. J. Natl. Cancer Inst. 67: 1353-1362, 1981. PubMed: 6273638
