| Applications |
PgsC-605 is a Chinese hamster ovary cell mutant deficient in sulfate transporter. The cell line was derived from CHO-K1 cells (see ATCC CCL-61) treated with mutagen (ethylmethanesulfonate) and isolated by replica plating [35S]sulfate colony autoradiography. PgsC-605 cells continue to produce heparin sulfate and chondroitin sulfate chains, but depend on endogenous formation of sulfate from sulfur containing amino acids. |
| Comments |
PgsC-605 is a Chinese hamster ovary cell mutant deficient in sulfate transporter. The cell line was derived from CHO-K1 cells (see ATCC CCL-61) treated with mutagen (ethylmethanesulfonate) and isolated by replica plating [35S]sulfate colony autoradiography. PgsC-605 cells continue to produce heparin sulfate and chondroitin sulfate chains, but depend on endogenous formation of sulfate from sulfur containing amino acids. |
| Subculturing |
Volumes used in this protocol are for 75 cm2 flask; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.
- Remove and discard culture medium.
- Briefly rinse the cell layer with 0.25% (w/v) Trypsin-0.53mM 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.
- Add appropriate aliquots of the cell suspension to new culture vessels.
- Incubate cultures at 37°C.
Subcultivation Ratio: 1:4 to 1:8
Medium Renewal: Every 2 to 3 days
Note: For more information on enzymatic dissociation and subculturing of cell lines consult Chapter 10 in Culture of Animal Cells, a Manual of Basic Technique by R. Ian Freshney, 3rd edition, published by Alan R. Liss, N.Y., 1994. |
| References |
Esko JD, et al. Tumor formation dependent on proteoglycan biosynthesis. Science 241: 1092-1096, 1988. PubMed: 3137658
Esko JD, et al. Sulfate transport-deficient mutants of Chinese hamster ovary cells. Sulfation of glycosaminoglycans dependent on cysteine. J. Biol. Chem. 261: 15725-15733, 1986. PubMed: 3782085
Elgavish A, et al. Chinese hamster ovary cell mutants deficient in an anion exchanger functionally similar to the erythroid band 3. J. Biol. Chem. 263: 18607-18613, 1988. PubMed: 3198592
Hay, R. J., Caputo, J. L., and Macy, M. L., Eds. (1992), ATCC Quality Control Methods for Cell Lines. 2nd edition, Published by ATCC.
Caputo, J. L., Biosafety procedures in cell culture. J. Tissue Culture Methods 11:223-227, 1988.
Fleming, D.O., Richardson, J. H., Tulis, J.J. and Vesley, D., (1995) Laboratory Safety: Principles and Practice. Second edition, ASM press, Washington, DC.
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