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BT-474 Clone 5; Breast Ductal Carcinoma; Human (Homo sapiens) Item ID: ATCCRL3247

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Organism Homo sapiens, human
Tissue mammary gland; breast/duct
Product Format frozen
Morphology epithelial-like
Culture Properties adherent, patchy (The cells form adherent patches of epithelial-like cells. The patches are compact multilayered colonies that rarely become confluent)
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 ductal carcinoma
Age 60 years
Gender female
Ethnicity Caucasian
Applications This cell line is valuable in the study of Her2 positive breast cancer resistant to herceptin therapy
Storage Conditions liquid nitrogen vapor phase
Images
Derivation BT-474 (ATCC HTB-20) cells were grown in 10ug/mL Herceptin for several weeks until the resulting cells (clones) were defined as resistant to Herceptin treatment. These resistant cells were grown in standard RPMI with 10% fetal bovine serum for many years under different conditions but the cells enclosed have not been exposed to Herceptin after they were first treated. They were not grown with Herceptin to make sure they stayed resistant. However, these cells have remained resistant for 30+ passages and they still retain the overexpression of HER-2 and are resistant to growth inhibition due to Herceptin.
Clinical Data 60 years adult Caucasian female
Receptor Expression Her2, expressed
Comments BT-474 (HTB-20) cells were grown in 10 µg/mL Herceptin for several weeks until the resulting cells (clones) were defined as resistant to Herceptin treatment. These resistant cells were grown in standard RPMI with 10% fetal bovine serum for many years under different conditions but the cells enclosed have not been exposed to Herceptin after they were first treated. They were not grown with Herceptin to make sure they stayed resistant. However, these cells have remained resistant for 30+ passages and they still retain the overexpression of HER-2 and are resistant to growth inhibition due to Herceptin. This cell line has overexpressed HER-2 and yet is resistant to herceptin (Trastuzumab) under in vitro conditions but sensitive to ADCC activity.
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 are given for a 75 cm2 flask. Increase or decrease the amount of dissociation medium needed proportionally for culture vessels of other sizes.
  1. Remove and discard culture medium.
  2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin- 0.53 mM 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.
Subcultivation Ratio: A subcultivation ratio of 1:2 is recommended Medium Renewal: 2 to 3 times per week
Cryopreservation Freeze medium: Fetal Bovine Serum, 92.5%; DMSO, 7.5% Storage temperature: liquid nitrogen vapor phase
Culture Conditions Temperature: 37°C Atmosphere: air, 95%; carbon dioxide (CO2), 5%
Permits

These permits may be required for shipping this product to Australia:

  • DAFF Import Permit formerly known as AQIS Import Permit must be obtained and a copy of the permit must be sent to ATCC in advance of shipment.
Basic Documentation Product Sheet Certificate of Analysis SDS
Other Documentation Cell Micrograph
References

Kute T, et al. Development of herceptin resistance in breast cancer cells. Cytometry A 57(2): 86-93, 2004. PubMed: 14750129

Kute T, et al. Understanding key assay parameters that affect measurements of trastuzumab-mediated ADCC against Her2 positive breast cancer cells. Oncoimmunology 1(6): 810-821, 2012. PubMed: 23162748

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