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CT26.WT; Colon Carcinoma; Mouse (Mus musculus) Item ID: ATCCRL2638

Organism Mus musculus, mouse
Tissue colon
Cell Type fibroblast
Product Format frozen
Morphology fibroblast
Culture Properties adherent
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.
Disease carcinoma
Strain BALB/c
Applications The cell line can be used with CT26.CL25 (ATCC CRL-2639) as a model for testing immunotherapy protocols and in studies on the host immune response.
Storage Conditions liquid nitrogen vapor phase
Derivation CT26 is an N-nitroso-N-methylurethane-(NNMU) induced, undifferentiated colon carcinoma cell line. It was cloned to generate the cell line designated CT26.WT (ATCC CRL-2638).
Antigen Expression H-2d RefWang M, et al. Active immunotherapy of cancer with a nonreplicating recombinant fowlpox virus encoding a model tumor-associated antigen. J. Immunol. 154: 4685-4692, 1995. PubMed: 7722321
Tumorigenic Yes
Effects Yes, in BALB/c mice. Mice inoculated, subcutaneously, developed lethal tumors at 80% frequency with 10(3) cells and at 100% with 10(4) cells. Pulmonary metastases developed when mice were inoculated, intravenously, with 10(4) cells. RefWang M, et al. Active immunotherapy of cancer with a nonreplicating recombinant fowlpox virus encoding a model tumor-associated antigen. J. Immunol. 154: 4685-4692, 1995. PubMed: 7722321
Comments CT26.WT was stably transduced with the retroviral vector LXSN that contains the lacZ gene encoding the model tumor associated antigen (TAA), beta-galactosidase (beta-gal) to obtain the lethal subclone CT26.CL25 (ATCC CRL-2639).

The growth rate and lethality of CT26.CL25 and CT26.WT is virtually identical despite the expression by CT26.CL25 of the model TAA, beta-galactosidase, in normal mice.

A culture submitted to the ATCC in July of 2001 was found to be contaminated with mycoplasma. Progeny were cured by a 21-day treatment with BM Cycline.

The cells were assayed for mycoplasma, by the Hoechst stain, PCR and the standard culture test, after a six-week period following treatment. All tests were negative.
Complete Growth Medium The base medium for this cell line is ATCC-formulated RPMI-1640 Medium, ATCC 30-2001. To make the complete growth medium, add the following components to the base medium: fetal bovine serum (ATCC 30-2020) to a final concentration of 10%.
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.
  1. Remove and discard culture medium.
  2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin-0.03% (w/v) EDTA solution to remove all traces of serum that contains trypsin inhibitor.
  3. 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.
  4. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
  5. Add appropriate aliquots of the cell suspension to new culture vessels.
  6. Incubate cultures at 37°C
Subculture ratio: 1:4 to 1:10 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
Cryopreservation Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO Storage temperature: liquid nitrogen vapor phase
Culture Conditions Atmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37°C
Name of Depositor N Restifo
References

Wang M, et al. Active immunotherapy of cancer with a nonreplicating recombinant fowlpox virus encoding a model tumor-associated antigen. J. Immunol. 154: 4685-4692, 1995. PubMed: 7722321

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