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Permits and Restrictions	View Permits
Organism	Mus musculus, mouse
Tissue	breast epithelium (mammary gland)
Cell Type	epithelial cell
Product Format	frozen
Morphology	epithelial-like
Culture Properties	adherent
Biosafety Level	2 [Cells contain CMV and SV40 viral DNA sequences] 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.
Gender	female
Applications	This cell line can be used to study normal epithelial cell function and cell transformation.
Storage Conditions	liquid nitrogen vapor phase

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Derivation	EpH4-Ev was produced by transfecting parental EpH4, a mouse mammary epithelial cell line, with an empty expression vector carrying a puromycin resistance gene. Stable clones were isolated by selection in medium containing 1.2 µg/mL puromycin.
Genes Expressed	puromycin resistance gene, expressed
Tumorigenic	No
Comments	EpH4-Ev was produced by transfecting parental EpH4, an immortalized mouse mammary epithelial cell line, with an empty expression vector carrying a puromycin resistance gene. Stable clones were isolated by selection in medium containing 1.2 mg/mL puromycin. The EpH4 cell line presents an attractive model in which to study transformation because the cells exhibit morphological and functional characteristics typical of normal mammary epithelial cells. This cell line has been used to generate constitutively activated MEK1 transformed cell line B-MEKDD 116 (ATCC CRL-3069). There are three additional related cell lines: EpH 1424 (ATCC CRL-3071), EpH4 1424.1 (ATCC CRL-3209), EpH4 1424.2 (ATCC CRL-3210).

Complete Growth Medium	The base medium for this cell line is ATCC-formulated DMEM Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium: 10% Bovine Calf Serum 1.2 mcg/mL Puromycin
Subculturing	Volumes used in this protocol are for 75 cm² flasks; 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 Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS) or 0.25% (w/v) Trypsin - 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor. Add 1.0 to 2.0 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. An inoculum of 4 x 10³ to 8 x 10³ viable cells/cm² is recommended. Incubate cultures at 37°C. Subculture when the cell concentration is between 8 x 10⁴ to 1.5 x 10⁵ cells/cm². Subcultivation ratio: A subcultivation ratio of 1:6 to 1:15 is recommended.Medium renewal: Every 2 to 3 days
Cryopreservation	Freeze medium: complete growth medium supplemented with an additional 10% bovine calf serum and 10% DMSOStorage temperature: liquid nitrogen vapor phase
Culture Conditions	Temperature: 37°CAtmosphere: air, 95%; carbon dioxide (CO₂), 5%

Name of Depositor	P Leder
References	Fantin VR, et al. A novel mitochondriotoxic small molecule that selectively inhibits tumor cell growth. Cancer Cell 2(1): 29-42, 2002 PubMed: 12150823

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