HeLa, U2OS, and HEK293T cells were cultured in Dulbecco's Modified Eagle Medium (DMEM) with 10% fetal bovine serum, 100 U/ml penicillin, and 100 μg/ml streptomycin in a humidified incubator containing 5% CO2 at 37 °C. DMEM/Nutrient Mixture F-12 medium (DMEM/F-12) was used for culturing nontransformed RPE cells, as previously described (7 (link)). UWB 1.289 cells and UWB 1.289 cells reconstituted with BRCA1 (gifts of Dr Alessandro Vindigni) were cultured at 37 °C with 5% CO2 in 50% RPMI media, 50% Mammary Epithelial Cell Growth Medium BulletKit (Lonza CC-3150) supplemented with 3% fetal bovine serum, 100 U/ml penicillin, and 100 μg/ml streptomycin.
Mammary epithelial cell growth medium bulletkit
Manufactured by Lonza
Sourced in United States, Switzerland
The Mammary Epithelial Cell Growth Medium BulletKit is a complete, ready-to-use medium formulated for the optimal growth and maintenance of human mammary epithelial cells in vitro. It contains the necessary growth factors, hormones, and supplements required for the culture of these cell types.
Automatically generated - may contain errors
Lab products found in correlation
Mcf 10a, by American Type Culture Collection (5 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Mcf 10a, by Lonza (3 mentions)
Skbr3, by American Type Culture Collection (3 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Cholera toxin, by Merck Group (2 mentions)
Hs578t, by American Type Culture Collection (2 mentions)
Mcf 7, by Thermo Fisher Scientific (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Mccoy s 5a medium, by Thermo Fisher Scientific (2 mentions)
Mda mb 231, by American Type Culture Collection (2 mentions)
Htert hme1, by American Type Culture Collection (1 mentions)
Bt 549, by Thermo Fisher Scientific (1 mentions)
Minimum essential medium (mem), by Thermo Fisher Scientific (1 mentions)
Sk br 3, by Thermo Fisher Scientific (1 mentions)
Leibovitz s l 15 medium, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle medium, by Thermo Fisher Scientific (1 mentions)
Mda mb 231, by Thermo Fisher Scientific (1 mentions)
Py8119, by American Type Culture Collection (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
10 protocols using mammary epithelial cell growth medium bulletkit
1
Culturing Diverse Cell Lines for Research
2
Breast Cancer Cell Line Characterization
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Human HER2+ BT474 (HTB-30) and SKBR3 (HTB-20) BC cell lines were acquired from ATCC. The brain-seeking subline of the triple-negative MDA-MB-231 cells, MDA-MB-231/BR, was a generous gift of Dr. Patricia Steeg (NCI, Bethesda, Maryland). A patient-derived HER2+ BCBM94 BC cell line was established in the laboratory from BC cells isolated from a surgically resected HER2+ BC brain metastasis. The first mouse brain passage of the BCBM94 model was used in the study. All cell lines were routinely cultured in DMEM/F-12 Ham’s medium supplemented with 10% FBS. Except for data presented in Fig. 6A –C in which 10 % FBS was supplemented during the assay, all treatments were done in DMEM/F-12 Ham’s 1% FBS. hTERT-HME1 (CRL-4010) were acquired from ATCC and cultured in Mammary Epithelial Cell Growth Medium Bullet Kit from Lonza (Bend, OR, Catalog #: CC-3150).
3
Culturing and Maintaining Human Breast Cell Lines
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Human breast cells MCF-10A, MCF-7, MDA-MB-231, SK-BR-3 and BT549 were purchased from the American Type Culture Collection (ATCC). The murine breast cancer PY8119 cell line was purchased from ATCC. MCF-10A cells were cultured in Mammary Epithelial Cell Growth Medium BulletKit (Lonza/Clonetics Corporation). MCF-7 cells were maintained in Minimum Essential Medium (Gibco) supplemented with 10% fetal bovine serum (FBS) (Gibco) and 10 μg/ml insulin (Sigma). MDA-MB-231 cells were cultured in Leibovitz’s L-15 Medium (Gibco) supplemented with 10% FBS. SK-BR-3 cells were cultured in McCoy’s 5a Medium (Gibco) supplemented with 10% FBS, whereas BT549 cells were maintained in RPMI-1640 (Gibco) containing 10% FBS. The PY8119 cell line was cultured in F12K (HyClone) supplemented with 5% FBS. HEK293T cells from ATCC were cultured in dulbecco’s modified eagle medium (Gibco) containing 10% FBS. Incubate the MDA-MB-231 cells at 37 °C incubator without CO2. Other cells were maintained at 37 °C, 5% CO2. Anti-mycoplasma reagent SaveIt (Hanbio) and 0.1% penicillinstreptomycin (Thermo) were used in all cell culture system. All cell lines were authenticated by STR profiling. Petri dishes and cell culture plates were purchased from Jet Bio-Filtration Co., Ltd (Guangzhou, China) and NEST Biotechnology Co. Ltd. (Wuxi, China).
4
Breast Cancer Cell Lines Culturing
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Human breast carcinoma cell line (HS578T) and mammary breast fibrocystic disease cell line (MCF10a), obtained from the American Type Tissue Collection (Rockville, MD, USA), were grown in DMEM (GIBCO) and MEGM (Lonza, Basel, Switzerland), respectively. DMEM was supplemented with 10% heat inactivated FBS (GIBCO), 100 U/mL penicillin, 100 mg/mL streptomycin, and 1% L-glutamine. MEGM was supplemented with Mammary Epithelial Cell Growth Medium Bullet Kit (Lonza), 100 nM cholera toxin (Sigma-Aldrich) and 5% heat inactivated FHS (Lonza). All cell lines were grown at 37 °C in a 5% CO2 atmosphere.
5
Enrichment of Extracellular Vesicles from NCI-60 Cell Lines
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Sixty cell lines from the National Cancer Institute (NCI-60) were acquired and cultured, as previously described [40 (link)]. MCF10a cells were grown using the Mammary Epithelial Cell Growth Medium BulletKit (Lonza, Basel, Switzerland, CC-3150) comprised of the basal medium MEBM supplemented with the provided aliquots of bovine pituitary extract (BPE), human recombinant epidermal growth factor (hEGF), insulin, and hydrocortisone. Instead of the GA-1000 aliquot provided in the kit, 100 ng/mL of cholera toxin was added to the medium, as recommended by the American Type Culture Collection (ATCC). At 90% confluence, the complete medium was aspirated and cells were washed with warm sterile phosphate buffered saline (PBS). To minimize contaminating proteins, cells were grown in BPE-free medium for another 48 h before EV enrichment.
6
Breast Cancer Cell Line Cultivation
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MCF10A cells (ATCC) were grown in mammary epithelial cell growth medium bullet kit (Lonza). MCF7 cells (ATCC), MDA231 cells (ATCC) and SKBR3 cells (ATCC) were grown in Dulbecco’s Modified Eagle’s medium (DMEM) (Sigma) supplemented with 10% foetal calf serum (FCS), 2 mM L-glutamine (PAA Laboratories) and 1% penicillin/streptomycin (penicillin 10,000 units/ml; streptomycin 10 mg/ml) (BioSera). Cells were grown at 37°C, 5% CO2 in a sterile humidified incubator and passaged at ≥80% confluence. MCF10A is a non-malignant breast cell line and MCF7 is a luminal A breast cancer cell line classified as ER+, PR+/- and HER2-. MDA231 is a basal-like triple negative breast cancer cell line classified as ER-, PR- and HER2- and SKBR3 is a HER2+ cell line classified as ER-, PR- and HER2+.
7
Culturing Breast Cell Lines for Research
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Human breast carcinoma cell line (HS578T) and mammary breast fibrocystic disease cell line (MCF10a), obtained from the American Type Tissue Collection (Rockville, MD, United States), were grown in DMEM (GIBCO) and MEGM (Lonza), respectively. DMEM was supplemented with 10% heat inactivated FBS (GIBCO), 100 U/ml penicillin, 100 mg/ml streptomycin, and 1% L-glutamine. MEGM was supplemented with Mammary Epithelial Cell Growth Medium Bullet Kit (Lonza),100 nM cholera toxin (Sigma Aldrich) and 5% heat inactivated FHS (Lonza). All cell lines were grown at 37°C in a 5% CO2 atmosphere.
8
Cell Line Cultivation and Maintenance
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Triple-negative, claudin-low BCCs MDA-MB-231, normal breast epithelial cells MCF-10A, and non–small cell lung cancer cells H1299 were purchased from ATCC. MDA-MB-231 and H1299 were cultured in DMEM supplemented with 10% FBS and 1% penicillin–streptomycin (herein defined as cancer medium). MCF-10A were cultured in Mammary Epithelial Cell Growth Medium BulletKit (Lonza).
9
Culturing Breast Cancer Cell Lines
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The breast cancer epithelial cell line MCF-10A (American Type Culture Collection [ATCC], Manassas, VA, USA) was cultured in 100 ng/mL cholera toxin-supplemented Mammary Epithelial Cell Growth Medium BulletKitTM (Lonza/Clonetics Corporation, Walkersville, MD, USA). The breast cancer cell lines MCF-7 and MDA-MB-468 were obtained from the Chinese Academy of Medical Sciences (Shanghai, China). MCF-7 cells were maintained in minimum essential medium supplemented with 10% fetal bovine serum ([FBS]; Gibco). The breast cancer cell lines MDA-MB-231 and SK-BR-3 (ATCC) were cultured in L-15 and McCoy’s 5A medium (Gibco), both of which were supplemented with 10% FBS. The culture conditions for MDA-MB-468 cells were the same as for MDA-MB-231 cells. All cells were cultured at 37°C in saturated humidity under 5% CO2 atmosphere.
10
Cell Culture and Transfection Protocols
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Human fibrosarcoma cells (HT1080), MDA-MB-231, and PC-3 were obtained from the Korean Cell Line Bank (Seoul, Republic of Korea). Cells were maintained in Dulbecco's modified Eagle's medium and RPMI 1640 medium supplemented with 10% fetal bovine serum and 100 units/mL penicillin/streptomycin. MCF10A, and RWPE-1 cells were obtained from the ATCC (American Type Culture Collection, VA, USA). MCF10A was maintained in Mammary Epithelial Cell Growth Medium BulletKitTM (Lonza, Basel, Switzerland) with 100 ng/mL cholera toxin. RWPE-1 cell was maintained in Keratinocyte SFM (Gibco). All cells were cultured at 37 °C in 5% CO2 incubator. Cell lines were routinely tested for the absence of Mycoplasma by PCR using e-Myco™ plus Mycoplasma PCR Detection Kit (Intron Biotechnology, Korea). Cell transfection was performed using Lipofectamine 3000 (Invitrogen) and Viromer® RED (Lipocalyx, Halle, Germany) according to the manufacturer's instructions. For pC1-Hyper-Reder HT1080 stable cell line, cells were transfected and selected with 500 μg/mL G418 (Geneticin, Gibco) for 2 weeks and maintained with 100 μg/mL G418.
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