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.

This cell line is an excellent tool for research into aberrant cytosine methylation of CpG island sites and BRCA1 repression in sporadic breast cancer. It is also very useful as a methylated BRCA1 control.

UACC-3199 was derived from a 58 year-old female with infiltrating ductal carcinoma of the breast metastatic to the left axillary lymph nodes.xa0

female

The patient had a family history of breast cancer.

58 years old

Caucasian

epidermal growth factor receptor (EGF), expressed

estrogen receptor, not expressed

progesterone receptor, not expressed

c-erbB-2 positive

Volumes used in this protocol are for 75 cm² flasks; 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 Ca++/Mg++ free Dulbeccos 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.
3. 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.
4. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
5. Transfer cell suspension to a centrifuge tube and spin at approximately 125 x g for 5 to 10 minutes. Discard supernatant.
6. Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 1 X 10⁴ to 2 X 10⁴ viable cells/cm² is recommended.
7. Incubate cultures at 37°C. Subculture when the cell concentration is between 2 X 10⁴ to 5 X 10⁴ cells/cm².

Subcultivation ratio: A subcultivation ratio of 1:2 to 1:3 is recommended

Medium renewal: Every 3 to 4 days

Growth Conditions:xa0Cells grow very slowly. Cultures reach confluence in 2 to 3 weeks. Avoid inoculating vessels below recommended seeding density.

Freeze medium: complete growth medium, 90%; DMSO, 10%

Storage temperature: liquid nitrogen vapor phase

Temperature: 37°C

Atmosphere: air, 100%

CSF1PO: 12

D13S317: 11

D16S539: 12

D5S818: 13

D7S820: 10

THO1: 9.3

TPOX: 10, 13

vWA: 15, 17

Amelogenin: X