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Volumes used in this protocol are for 75 cm² flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes. Corning^® T-75 flasks (catalog #430641) are recommended for subculturing this product.

Note: if floating cells are present, it is recommended that they be transferred at the first two (2) subcultures as described below. It is not necessary to transfer floating cells for subsequent subcultures.

1. Remove culture medium to a centrifuge tube.
2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin - 0.53 mM EDTA solution to remove all traces of serum which 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. Transfer the cell suspension to the centrifuge tube with the medium and cells from step 1, and centrifuge at approximately 125 xg for 5 to 10 minutes. Discard the supernatant.
6. Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels.
7. Incubate cultures at 37°C.

Sugarman BJ, et al. Recombinant human tumor necrosis factor-alpha: effects on proliferation of normal and transformed cells in vitro. Science 230: 943-945, 1985. PubMed: 3933111

Takahashi K, Suzuki K. Association of insulin-like growth-factor-I-induced DNA synthesis with phosphorylation and nuclear exclusion of p53 in human breast cancer MCF-7 cells. Int. J. Cancer 55: 453-458, 1993. PubMed: 8375929

Brandes LJ, Hermonat MW. Receptor status and subsequent sensitivity of subclones of MCF-7 human breast cancer cells surviving exposure to diethylstilbestrol. Cancer Res. 43: 2831-2835, 1983. PubMed: 6850594

Lan MS, et al. Polypeptide core of a human pancreatic tumor mucin antigen. Cancer Res. 50: 2997-3001, 1990. PubMed: 2334903

Pratt SE, Pollak MN. Estrogen and antiestrogen modulation of MCF7 human breast cancer cell proliferation is associated with specific alterations in accumulation of insulin-like growth factor-binding proteins in conditioned media. Cancer Res. 53: 5193-5198, 1993. PubMed: 7693333

Huguet EL, et al. Differential expression of human Wnt genes 2, 3, 4, and 7B in human breast cell lines and normal and disease states of human breast tissue. Cancer Res. 54: 2615-2621, 1994. PubMed: 8168088

Soule HD, et al. A human cell line from a pleural effusion derived from a breast carcinoma. J. Natl. Cancer Inst. 51: 1409-1416, 1973. PubMed: 4357757

Bellet D, et al. Malignant transformation of nontrophoblastic cells is associated with the expression of chorionic gonadotropin beta genes normally transcribed in trophoblastic cells. Cancer Res. 57: 516-523, 1997. PubMed: 9012484

Littlewood-Evans AJ, et al. The osteoclast-associated protease cathepsin K is expressed in human breast carcinoma. Cancer Res. 57: 5386-5390, 1997. PubMed: 9393764

Komarova EA, et al. Intracellular localization of p53 tumor suppressor protein in gamma-irradiated cells is cell cycle regulated and determined by the nucleus. Cancer Res. 57: 5217-5220, 1997. PubMed: 9393737

van Dijk MA, et al. A functional assay in yeas for the human estrogen receptor displays wild-type and variant estrogen receptor messenger RNAs present in breast carcinoma. Cancer Res. 57: 3478-3485, 1997. PubMed: 9270016

Landers JE, et al. Translational enhancement of mdm2 oncogene expression in human tumor cells containing a stabilized wild-type p53 protein. Cancer Res. 57: 3562-3568, 1997. PubMed: 9270029

Umekita Y, et al. Human prostate tumor growth in athymic mice: inhibition by androgens and stimulation by finasteride. Proc. Natl. Acad. Sci. USA 93: 11802-11807, 1996. PubMed: 8876218

Zamora-Leon SP, et al. Expression of the fructose transporter GLUT5 in human breast cancer. Proc. Natl. Acad. Sci. USA 93: 1847-1852, 1996. PubMed: 8700847

Geiger T, et al. Antitumor activity of a PKC-alpha antisense oligonucleotide in combination with standard chemotherapeutic agents against various human tumors transplanted into nude mice. Anticancer Drug Des. 13: 35-45, 1998. PubMed: 9474241

Jang SI, et al. Activator protein 1 activity is involved in the regulation of the cell type-specific expression from the proximal promoter of the human profilaggrin gene. J. Biol. Chem. 271: 24105-24114, 1996. PubMed: 8798649

Lee JH, et al. The proximal promoter of the human transglutaminase 3 gene. J. Biol. Chem. 271: 4561-4568, 1996. PubMed: 8626812

Chang K, Pastan I. Molecular cloning of mesothelin, a differentiation antigen present on mesothelium, mesotheliomas, and ovarian cancers. Proc. Natl. Acad. Sci. USA 93: 136-140, 1996. PubMed: 8552591

Zhu X, et al. Cell cycle-dependent modulation of telomerase activity in tumor cells. Proc. Natl. Acad. Sci. USA 93: 6091-6095, 1996. PubMed: 8650224

Bacus SS, et al. Differentiation of cultured human breast cancer cells (AU-565 and MCF-7) associated with loss of cell surface HER-2/neu antigen. Mol. Carcinog. 3: 350-362, 1990. PubMed: 1980588

Huguet EL, et al. Differential expression of human Wnt genes 2, 3, 4, and 7B in human breast cell lines and normal and disease states of human breast tissue. Cancer Res. 54: 2615-2621, 1994. PubMed: 8168088

Nucleotide (GenBank) : U26553 Human calcitonin receptor mRNA, complete cds.

Nucleotide (GenBank) : U26554 Human calcitonin receptor isoform mRNA, complete cds.

Nucleotide (GenBank) : U63917 Human G protein coupled receptor (GPCR-Br) mRNA, complete cds.