| Comments |
The SR-4987 cell line was established by treating normal marrow cells with supernatant from Y-1 cells (known to produce Murine Leukemia Virus (MuLV). The cells produce MCSF, and are highly sensitive to fibroblast growth factor (FGF). They are poorly clonogenic in soft agar. Neither interleukin 3 (IL-3) or granulocyte colony stimulating factor (GCSF) were detected in SR-4987 conditioned medium. |
| 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 |
Pessina A, et al. Establishment and characterization of a new murine cell line (SR-4987) derived from marrow stromal cells. Cytotechnology 8: 93-102, 1992. PubMed: 1382506
Pessina A, et al. Expression of B cell markers on SR-4987 cells derived from murine bone marrow stroma. Exp. Hematol. 25: 536-541, 1997. PubMed: 9197333
Pessina A, et al. Role of SR-4987 stromal cells in the modulation of doxorubicin toxicity to in vitro granulocyte-macrophage progenitors (CFU-GM). Life Sci. 65: 513-523, 1999. PubMed: 10462078
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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