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微信小章| 产品名称： | CTLL-2 |
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| 商品货号： | TS212308 |
| Organism： | Mus musculus, mouse |
| Cell Type： | lymphocyte cytotoxic T lymphocyte; |
| Product Format： | frozen |
| Morphology： | lymphoblast |
| Culture Properties： | suspension |
| Biosafety Level： | 1
Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country. |
| Strain： | C57BL/6 |
| Applications： | The cells are dependent upon IL-2 for growth, and can be used to assay for IL-2. |
| Storage Conditions： | liquid nitrogen vapor phase |
| Images： |  |
| Derivation： | This line is a clone of cytotoxic T cells derived from a C57BL/6 mouse. |
| Receptor Expression： | interleukin 2 (IL-2), expressed |
| Comments： | The cells are dependent upon IL-2 for growth, and can be used to assay for IL-2. Immediately after thawing (or after overgrowth of a culture) the culture will appear to have few or no viable cells; this is normal. Depending upon the source and potency of the IL-2, it may take up to three weeks before the cells are ready to subculture. T-STIM with Con A (rat IL-2 culture supplement from Becton Dickinson) may be used or the rat factor may be prepared as described below. The freshly prepared rat factor will usually produce more rapid growth. Tested and found negative for ectromelia virus (mousepox). T-STIM with Con A (rat IL-2 culture supplement from Becton Dickinson) may be used or the rat factor may be prepared as described on the product sheet. The freshly prepared rat factor will usually produce more rapid growth. Rat Growth Factorxa0 (Laboratory Preparation)
Spleens are removed from female Sprague-Dawley rats weighing 200 g. They are coarsely minced before passing through a No. 60 sieve. Wash cells 2-3 times with RPMI 1640. Resuspend cells at 1-1.5 X 106xa0 viable cells/ml with 100-200 mL/150 cm2 flask in RPMI 1640 containing 1%xa0 heat-inactivated fetal bovine serum, 0.05 mM,xa0 2-mercaptoethanol, 15 mM HEPES, 100 units/mL penicillin, 100 mg/mL streptomycin and 1.0 µg/mL concanavalin A.xa0xa0xa0xa0xa0xa0xa0xa0xa0
Incubate at 37°C in a CO2 incubator for 48 hours. Harvest the supernatant by centrifugation at 16,000 x g for 10 minutes at 4°C. Sterilize by filtrationxa0 using a 0.22 micron membrane.
Store at -60°C. Avoid freeze-thaw cycles. Rat growth factor stored at 4°C up to one month has retained its quality. The growth factor is added to the medium just before use. Expect 1-2 X 108 cells per spleen which yields 100-200 mL of growth factor.
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| Complete Growth Medium： | The base medium for this cell line is ATCC-formulated RPMI-1640 Medium, (ATCC®xa030-2001™). To make the complete growth medium, add the following components to the base medium: additional 2 mM L-glutamine; additional 1mM sodium pyruvate; adjust to a final concentration of 10% fetal bovine serum and 10% T-STIM with Con A. T-STIM is available from Becton Dickinson. |
| Subculturing： | Subculture actively growing suspension cultures before they have reached 2 X 105xa0cells/ml or the IL-2 will rapidly deplete and the cells will quickly lose viability Use inoculation densities of 1 to 2 X 104xa0viable cells/mL.xa0Corning® T-75 flasks (catalog #431464) are recommended for subculturing this product.
Medium Renewal:xa0Twice per week
Some Important Considerations in Handling CTLL-2, TIB-214xa0 Frozen Cells: Viability immediately after thawing will be 70-80%. Expect viability to be very poor from day 1 to day 4 after culture initiation. Culture will appear to be completely dead. On the third to fifth day following initiation viable cell clusters will begin to appear in suspension. Usually cells will be ready to subculture on the 7th to the 10th day after the ampule is thawed. However, it may take from 2-3 weeks before vigorous growth is observed. It is best to leave the initial culture undisturbed until cells enter their growth phase. Overgrowth: In the event cell density becomes too great and viability decreases to where culture appears totally dead, the culture may still be rescued. Inoculate a flask at a density of 1 X 104 viable cells/mL. |
| Cryopreservation： | Freeze medium: Complete growth medium supplemented with an additional 10% fetal bovine serum and 7.5% DMSO
Storage temperature: liquid nitrogen vapor phase |
| Culture Conditions： | Atmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37°C |
| Name of Depositor： | S Gillis |
| Deposited As： | Mus musculus |
| References： | Gillis S, Smith KA. Long term culture of tumour-specific cytotoxic T cells. Nature 268: 154-156, 1977. PubMed: 145543 Hu SX, et al. Development of an adenovirus vector with tetracycline-regulatable human tumor necrosis factor alpha gene expression. Cancer Res. 57: 3339-3343, 1997. PubMed: 9269991 Mazzaccaro RJ, et al. Major histocompatibility class I presentation of soluble antigen facilitated by Mycobacterium tuberculosis infection. Proc. Natl. Acad. Sci. USA 93: 11786-11791, 1996. PubMed: 8876215 Belani R, Weiner GJ. Expression of both B7-1 and CD28 contributes to the IL-2 responsiveness of CTLL-2 cells. Immunology 87: 271-274, 1996. PubMed: 8698390 Hay RJ, Caputo JL, Macy, ML, Eds. (1992) ATCC Quality Control Methods for Cell Lines. 2nd edition, Published by ATCC. Fleming, D.O., Richardson, J. H., Tulis, J.J. and Vesley, D., (1995) Laboratory Safety: Principles and Practice. Second edition, ASM press, Washington, DC. Caputo JL. Biosafety procedures in cell culture. J. Tissue Culture Methods 11:223-227, 1988 |
| Cross References： | Nucleotide (GenBank) : M27960 Mouse interleukin-4 receptor (secreted form) mRNA, complete cds. Nucleotide (GenBank) : M27959 Mouse (clone B-2) interleukin-4 receptor (membrane-bound form) mRNA, complete cds. |