Ct26 murine colorectal carcinoma

The CT26 murine colorectal carcinoma [85 (link)] (Experiment 1) and the Lewis lung carcinoma (3LL) murine cell line [86 (link)] (Experiments 2 and 3) were obtained from the ATCC (Manassas, VA, USA). Stocks were within 3 passages of the ATCC clones with in vitro expansion used to generate a bank of matched early passage cells for subsequent experiments. Cells were grown in RPMI-1640 media supplemented with HEPES, non-essential amino acids, sodium pyruvate, glutamine, 10% FBS, penicillin and streptomycin. All cell lines tested negative for mycoplasma by ELISA. To initiate tumor growth, cells were subcutaneously injected into the right flanks on experimental Day 0 at a dosage of 5 × 10⁵ of CT26 (Experiment 1) or 3LL (Experiments 2 and 3) cells suspended in 100 μL PBS per animal. Tumor-free mice were injected with PBS alone using the same procedure. In experiments 1 and 2, half of mice were tumor-free (TF) and half of mice were tumor-bearing (TB). In experiment 3, all mice received tumors.

The CT26 murine colorectal carcinoma [14 (link)] was obtained from ATCC (Manassas, VA). Cells were grown in RPMI-1640 media supplemented with HEPES, non-essential amino acids, sodium pyruvate, glutamine, 10% FBS, penicillin and streptomycin. All cell lines tested negative for mycoplasma. BALB/c and FVB mice were obtained from Jackson Laboratories (Bar Harbor, ME). FVB mice bearing the MMTV-PyMT transgene [15 (link)] were kindly provided by Dr. Akporiaye (EACRI, Portland OR) and heterozygous PyMT⁺ mice that spontaneously develop mammary tumors and PyMT^- tumor-free littermates were recruited into comparative studies. Tumor bearing mice were monitored a minimum of three days per week and euthanized when tumors exceeded 12mm in any dimension, or when body condition score declined one level. Euthanasia was performed with CO2 inhalation followed by a second method, either organ harvest or cervical dislocation. Radiation was performed with inhaled isoflurane anesthesia, intraperitoneal meloxicam was given for analgesia. There were no unexpected animal deaths. All animal protocols were approved by the Earle A. Chiles Research Institute IACUC (Animal Welfare Assurance No. A3913-01).