Bt483

Manufactured by American Type Culture Collection

Sourced in United States

The BT483 is a cell line derived from a human breast carcinoma. It is an adherent cell line that can be used for in vitro cell culture experiments. The core function of the BT483 cell line is to provide a model system for the study of breast cancer biology and the evaluation of potential therapeutic compounds.

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33 protocols using bt483

Cell Line Authentication and Maintenance

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BT483, MDA-MB-415, CAMA1, HCC1428, T47D, and MCF12A cells were obtained
from American Type Culture Collection (ATCC). ATCC distributed cell lines are
authenticated routinely through short tandem repeat (STR) profiling and are low
passage and contamination free. The MCF7-TamR and T47D-TamR cells were generated
in Dr. Rachel Schiff’s lab and were provided to us along with the
parental MCF7 cells, as previously described (18 (link)–20 (link)). The MCF7 cells
were authenticated by MD Anderson Cell Authentication Core Facility through STR
profiling. HCC1428, and BT483 cells were cultured in RPMI 1640 (ATCC) with
10%−20% fetal bovine serum and 200 mg/ml L-glutamine (Invitrogen)
according to ATCC recommendation. MDA-MB-415 cells were cultured in DMEM (Thermo
Fisher Scientific) with 10% fetal bovine serum and 200 mg/ml L-glutamine
(Invitrogen). MCF12A cells were cultured as previously described (21 (link)). Tamoxifen-resistant cells (TamR) were
established by continuous passage of parental cells in the presence of
10⁻⁷ M Tam. TamR cells were propagated in phenol red-free
RPMI 1640, with 5% charcoal-dextran treated fetal bovine serum (CD-FBS, Thermo
Fisher Scientific) and 10⁻⁷ M Tam. Cell lines are routinely
tested for mycoplasma contamination following a published protocol (13 (link)).

Wang X., Veeraraghavan J., Liu C.C., Cao X.X., Qin L., Kim J.A., Tan Y., Loo S.K., Hu Y., Lin L., Lee S., Shea M., Mitchell T., Li S., Ellis M., Hilsenbeck S.G., Schiff R, & Wang X.S. (2021). Therapeutic targeting of nemo-like kinase in primary and acquired endocrine-resistant breast cancer. Clinical cancer research : an official journal of the American Association for Cancer Research, 27(9), 2648-2662.

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Culturing Human Breast Cancer Cells

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Human breast cancer cell lines BT-483 and MCF-7 were purchased from ATCC (Manassas, USA). BT-483 cells were cultured with ATCC-formulated RPMI-1640 medium (cat. no. 30-2001; ATCC) containing 0.01 mg/mL bovine insulin and 20% fetal bovine serum (FBS). MCF-7 cells were cultured in ATCC-formulated Eagle's minimum essential medium (cat. no. 30-2003; ATCC) containing 0.01 mg/mL human recombinant insulin and 10% FBS. Cells were cultured at 37℃ and harvested during logarithmic growth phase for subsequent experiments.

Peng J., Wang X., Ran L., Song J., Luo R, & Wang Y. (2018). Hypoxia-Inducible Factor 1α Regulates the Transforming Growth Factor β1/SMAD Family Member 3 Pathway to Promote Breast Cancer Progression. Journal of Breast Cancer, 21(3), 259-266.

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Characterization of Breast Cancer Cell Lines

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The following cell lines were obtained from the American Type Culture Collection (ATCC): i) TNBC lines MDA-MB-157, MDA-MB-436, MDA-MB-468, HCC70, BT-549, ii) ER+ /HER2/neu negative cell lines T-47D, ZR-75-1, MCF-7, MDA-MB-415, HCC1428, BT-483, iii) ER⁺/ Her2/neu⁺ cell lines BT474, and MDA-MB-361 and iv) ER-/Her2/neu⁺ cell lines AU565, HCC1954, and SKBR3. All cells except HS578T were cultured in DMEM-F12 containing 10% FBS. The TNBC line, HS578T also obtained from ATCC, was grown in DMEM with reduced NaHCO₃ (ATCC) containing 0.1 mM insulin and 10% FBS. The ER⁺/ Her2/neu-overexpressing MCF7 (MCF7-Her18) cells were a kind gift from Dr. Elizabeth Mittendorf. All the cell lines used in here were strictly with in ten passages after buying from ATCC and thus were not authenticated again.

Bhardwaj A., Ganesan N., Tachibana K., Rajapakshe K., Albarracin C.T., Gunaratne P.H., Coarfa C, & Bedrosian I. (2015). Annexin A1 Preferentially Predicts Poor Prognosis of Basal-Like Breast Cancer Patients by Activating mTOR-S6 Signaling. PLoS ONE, 10(5), e0127678.

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Cell Line Characterization and Reagent Procurement

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The ZR75-1 and BT483 cell lines were obtained from the ATCC (Bethesda, MD). MCF7 cells derived from the original University of Michigan stock were kindly provided by Dr. K. Nephew (University of Indiana, Bloomington). Palbociclib and 5FU were purchased from Selleckchem (Houston, TX). All Materials were obtained as described in the references [1 (link)–5 (link)]. Trypsin-EDTA, DMEM, RPMI, penicillin-streptomycin were purchased from GIBCOBRL (GIBCOBRL Life Technologies, Grand Island, NY). Other reagents and performance of experimental procedures were as described [1 (link)–5 (link)]. Antibodies were purchased from Cell Signaling Technology (Danvers, MA); Abgent (San Diego, CA); Novus Biologicals (Centennial, CO); Abcam (Cambridge, UK); and Santa Cruz Biotechnology (Dallas, TX). Specific multiple independent siRNAs to knock down the expression of CD95, FADD, Beclin1, ATG5 AMPKa₁, ATM, BIM, BAX, BAK, BID and eIF2α, and scramble control, were purchased from Qiagen (Hilden Germany). Control studies were presented showing on-target specificity of our siRNAs, primary antibodies, and our phospho-specific antibodies to detect both total protein levels and phosphorylated levels of proteins [1 (link)–6 (link)] (Supplementary Figure 5).

Booth L., West C., Moore R.P., Von Hoff D, & Dent P. (2022). GZ17-6.02 and palbociclib interact to kill ER+ breast cancer cells. Oncotarget, 13, 92-104.

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Histogel Embedding of BT-483 Breast Cancer Cells

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Breast cancer cell lines MDA-MB-231, T-47D, AU565, and BT-483 were obtained from ATCC and SUM-185PE was obtained from Steve Ethier (University of Michigan). Cells were cultured in media recommended by the provider. The identity of the cell lines was confirmed by short tandem repeats analysis. Tests for mycoplasma contamination were performed regularly. For histogel embedding, BT-483 cells were fixed in 4% paraformaldehyde for 20 minutes, dehydrated in a series of ascending ethanol and resuspended in 2 volumes of histogel (Thermo Scientific), pre-warmed to 55°C. Once solidified, the histogel was kept in 70% ethanol and processed into paraffin block.

Janiszewska M., Liu L., Almendro V., Kuang Y., Paweletz C., Sakr R.A., Weigelt B., Hanker A.B., Chandarlapaty S., King T.A., Reis-Filho J.S., Arteaga C.L., Park S.Y., Michor F, & Polyak K. (2015). In situ single cell analysis identifies heterogeneity for PIK3CA mutation and HER2 amplification in HER2+ breast cancer. Nature genetics, 47(10), 1212-1219.

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Breast cancer cell line culture

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The cell lines used in this study: AU565, BT20, BT474, BT483, BT549, HCC1143, HCC1500, HCC1569, HCC1937, HCC1954, HCC38, HCC70, Hs578T, MDA-MB-134, MDA-MB-175, MDA-MB-361, MDA-MB-436, MDA-MB-453, MDA-MB-468, SKBR3, and ZR751 were obtained from ATCC, Rockville, MD, USA. MCF-7 cells were obtained from Michigan Cancer Foundation, Detroit, MI, USA. The cell lines HBL100, MDA-MB-157, MDA-MB-231 and T47D were obtained from EG&G Mason Research Institute, Worcester, MA, USA.
The cell lines AU565, BT20, BT474, BT549, HBL100, Hs578T, MCF-7, MDA-MB-134, MDA-MB-157, MDA-MB-175, MDA-MB-231, MDA-MB-361, MDA-MB-436, MDA-MB-453, MDA-MB-468, SKBR3, T47D and ZR571 were cultured in RPMI 1640 medium supplemented with 10% heat-inactivated fetal bovine serum (FBS), 6 mM L-glutamine, 20 mM HEPES and 10 μg/ml human insulin (CSL-Novo, North Rocks, NSW, Australia). The remaining cell lines were cultured in RPMI 1640 medium supplemented with 10% heat-inactivated FBS, 6 mM L-glutamine, 1 mM sodium pyruvate and 20 mM HEPES. The MYC over-expressing MCF7 cells have been previously described
[11 (link),27 (link)] and were cultured in the same conditions as the parental cells.
The CDK4/6 inhibitor PD0332991 was purchased from Selleck Chemicals (Houston, TX, USA), CDK2 inhibitor SNS-032 from Symansis (Auckland, New Zealand) and CDK1 inhibitors, RO-3306 and CGP74514A, from Calbiochem (San Diego, CA, USA).

Kang J., Sergio C.M., Sutherland R.L, & Musgrove E.A. (2014). Targeting cyclin-dependent kinase 1 (CDK1) but not CDK4/6 or CDK2 is selectively lethal to MYC-dependent human breast cancer cells. BMC Cancer, 14, 32.

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Mammary Gland Epithelial Adenocarcinoma Cell Lines

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Human mammary gland epithelial adenocarcinoma cell lines (luminal subtype A: BT474 (ATCC HTB-20), BT483 (ATCC HTB-121), MCF-7 (ATCC HTB-22), T47D (ATCC HTB-133), and ZR75-1 (ATCC CRL-1500) [42 (link)]; luminal subtype B: AU565 (ATCC CRL-2351), MDA-MB-453 (ATCC HTB-131), and SKBR3 (ATCC HTB-30) [43 (link)–45 (link)]; basal subtype: MDA-MB-231 (ATCC HTB-26) HS578T (ATCC HTB-126) [46 (link)]); the normal human breast epithelial cell line MCF-10A (ATCC CRL-10317); and an immortalized cell line obtained from a primary culture of cells derived from an early lactation sample of human milk, HBL100 (ATCC HTB-124), were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). MCF-10A cells were maintained in MCF-10A culture medium consisting of DMEM/F12 (Thermo Fisher Scientific, Passau, Germany) supplemented with 20 ng/mL epidermal growth factor, 10 g/mL insulin, 0.5 g/mL hydrocortisone, and 1X non-essential amino acids (Thermo Fisher Scientific). BT474, BT483, MCF-7, MDA-MB-453, MDA-MB-231, and HBL-100 cells were maintained in DMEM (Thermo Fisher Scientific). T47D, ZR-75 AU-565, SKBR3, and HS-578T cells were maintained in DMEM/F12 (Thermo Fisher Scientific). The cells were cultured according to standard protocols [21 (link)].

Tu S.H., Lin Y.C., Huang C.C., Yang P.S., Chang H.W., Chang C.H., Wu C.H., Chen L.C, & Ho Y.S. (2016). Protein phosphatase Mg2+/Mn2+ dependent 1F promotes smoking-induced breast cancer by inactivating phosphorylated-p53-induced signals. Oncotarget, 7(47), 77516-77531.

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Histogel Embedding of BT-483 Breast Cancer Cells

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Breast Cancer Cell Line Maintenance Protocol

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Human mammary gland epithelial adenocarcinoma cell lines (luminal subtype A: BT-483 (ATCC- HTB-121), MCF-7 (ATCC HTB-22), T47D (ATCC HTB-133), and ZR75-1 (ATC CRL-1500); luminal subtype B: BT-474 (ATCC HTB-20); HER2 subtype: AU-565 (ATCC CRL-2351), SKBR3 (ATCC HTB-30), HCC1419 (ATC CRL-2326), HCC1945 (ATC CRL-2338), and MDA-MB-453 (ATCC HTB-131); basal subtype: MDA-MB-231 (ATCC HTB-26), MDA-MB-468 (ATCC HTB-132), and Hs-578T (ATCC HTB-126)), a human melanoma cell line (MDA-MB-435s), and non-tumourigenic immortalized breast epithelial cell line [MCF-10A (ATCC CRL-10317)] were purchased from American Tissue Cell Culture collection (ATCC). MCF-10A cells were maintained under conditions that have been well-established by Xiao-guang Sun et al. [50]. The other cell lines were maintained in DMEM or DMEM/F12 (Thermo Fisher Scientific) supplemented with 10% FBS, Thermo Fisher Scientific), penicillin (100 units/mL), streptomycin (100 μM/mL), and 250 μg/mL amphotericin B at 37°C in a humidified 5% CO₂ incubator.

Lin J.H., Lee W.J., Wu H.C., Wu C.H., Chen L.C., Huang C.C., Chang H.L., Cheng T.C., Chang H.W., Ho C.T., Tu S.H, & Ho Y.S. (2019). Small G protein signalling modulator 2 (SGSM2) is involved in oestrogen receptor-positive breast cancer metastasis through enhancement of migratory cell adhesion via interaction with E-cadherin. Cell Adhesion & Migration, 13(1), 120-137.

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Comprehensive Cell Line Database

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All the cell lines were obtained from American Type Culture Collection (Manassas, USA), including human mammary epithelial (HME) cell lines (MCF10A and 184A1) and breast cancer cell lines (SKBR3, T47D, BT474, MCF-7, BT-483, BT-20, BT549, MDA-MB-468 and MDA-MB-231). All the cell lines were passaged in our laboratory for less than six months and maintained according to the supplier’s instructions. The cell lines were found to be free of mycoplasma infection and authenticity verified by DNA fingerprinting before use.

He R., Liu P., Xie X., Zhou Y., Liao Q., Xiong W., Li X., Li G., Zeng Z, & Tang H. (2017). circGFRA1 and GFRA1 act as ceRNAs in triple negative breast cancer by regulating miR-34a. Journal of Experimental & Clinical Cancer Research : CR, 36, 145.

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