HCT116 colon, MCF7 breast, MDA-MB-231 breast, A375 melanoma, and PC9 lung were purchased from ATCC, were they were validated. HCT116 AKT1/2−/− was purchased from Horizon Discovery (Cambridge, UK), where it was validated. AG11726 skin fibroblasts were purchased from Coriell Repositories, where they were validated. MCF7, MDA-MB-231 and AG11726 were maintained in DMEM, 10% FCS, 40mM glutamine, 100 U/mL penicillin, and 100μg/mL streptomycin; HCT116 and HCT116 AKT1/2−/− in McCoy’s 5α medium supplemented with 10% FCS, 100U/mL penicillin, and 100μg/mL streptomycin; PC9 in RPMI, 25% glucose, 1% sodium pyruvate, 100U/mL penicillin, and 100μg/mL streptomycin; A375 in DMEM supplemented with high glucose HEPES buffer, 10% FCS, 100U/mL penicillin, and 100μg/mL streptomycin. All the cells were grown at 37°C and 5% CO2.
Mcf 7 breast
Manufactured by American Type Culture Collection
Sourced in United States
MCF-7 is a human breast adenocarcinoma cell line. It is a commonly used model for breast cancer research.
Automatically generated - may contain errors
Lab products found in correlation
Mda mb 231, by American Type Culture Collection (3 mentions)
Hct116 colon, by American Type Culture Collection (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
U 251 glioblastoma gbm, by American Type Culture Collection (2 mentions)
Penicillin, by Thermo Fisher Scientific (2 mentions)
Ht29 colon cancer, by American Type Culture Collection (2 mentions)
L glutamine, by Thermo Fisher Scientific (2 mentions)
A375 melanoma, by American Type Culture Collection (1 mentions)
Mda mb 231 breast, by American Type Culture Collection (1 mentions)
Pc9 lung, by American Type Culture Collection (1 mentions)
Hct116 akt1 2, by Horizon Discovery (1 mentions)
Htb 72, by American Type Culture Collection (1 mentions)
B16f10 murine melanoma, by American Type Culture Collection (1 mentions)
Acc 107, by Leibniz Institute DSMZ (1 mentions)
Iscove s modified dulbecco s medium, by Merck Group (1 mentions)
Penicillin, by Merck Group (1 mentions)
Streptomycin, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by MP Biomedicals (1 mentions)
Du145, by American Type Culture Collection (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
18 protocols using mcf 7 breast
1
Cell Line Cultivation and Validation
2
Anticancer Activity Evaluation of Compounds
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Compounds 1 –8 were evaluated for their in vitro growth inhibitory activity against five human tumor cell lines, including the A549 non-small cell lung cancer (ACC107, Deutsche Sammlung von Mikroorganismen und Zellkulturen, Braunschweig, Germany), the U373 glioblastoma (ECACC 08061901, European Collection of Authenticated Cell Cultures, Salisbury, UK), the Hs683 oligodendroglioma (ATCC HTB-138, American Type Culture Collection, Manassas, VA, USA), the MCF7 breast cancer (ATCC HTB22) and the SKMEL28 melanoma (ATCC HTB72) cell lines, and against the B16F10 murine melanoma (ATCC CRL 6475) cell line. The inhibitory growth activity of the compounds under study was determined by means of the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] colorimetric assay [34 (link),35 (link)]. The cancer cells were cultured during three days in the presence of the compounds and the data were reported as mean IC50 values calculated on the sextuplicates of the experiment conducted once for each compound and for each cell line.
3
Characterization of B-cell Malignancy Cell Lines
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MCF7, breast adenocarcinoma cells (ATCC, Manassas, VA, USA) were grown in Dulbecco's modified Eagle's medium with 10% fetal bovine serum (Biomedicals, Wien, Austria), 100 U·mL−1 penicillin and 0.1 mg·mL−1 streptomycin (Sigma‐Aldrich, MERCK KGaA, Darmstadt, Germany). Cell lines representing B‐cell malignancies, SU‐DHL‐4 (diffuse large B‐cell lymphoma), WSU‐NHL (diffuse large B‐cell lymphoma) and MEC‐1 (chronic lymphocytic leukemia in prolymphocytoid transformation), were obtained from the Leibnitz Institute DSMZ‐German Collection of Microorganisms and Cell Cultures. These cells were cultured in Iscove's modified Dulbecco's medium from Sigma‐Aldrich supplemented with fetal bovine serum (MP Biomedicals, Wien, Austria). All cells were cultured in a humidified atmosphere at 37 °C and 5% CO2. The TP53 mutation status in cell lines from B‐cell malignancies was verified by yeast functional analysis (FASAY) coupled to sequencing 36 .
4
Cell Cycle Analysis of Cancer Cell Lines
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HU and propidium iodide (PI) were purchased from Sigma Aldrich (Oakville, ON). MCF7 breast cancer and DU145 prostate cancer cell lines were obtained from ATCC, and cultured in DMEM (MCF7) and MEM (DU145) supplemented with 10% FBS (Sigma Aldrich, Oakville, ON) and 1% Penicillin-Streptomycin (Life Technologies, Carlsbad, CA). Cell cycle distribution was examined according to our published procedure [71 (link)].
5
Cell Line Maintenance and Authentication
6
Establishment and Culture of Metastatic Breast Cancer Cell Lines
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MDA-MB-231 and MCF-7 breast tumour cells (ATCC, Manassas, VA, USA) were cultured in phenol-red free RPMI 1640 and DMEM respectively supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FCS), 2 mM L-glutamine, 1 mM nonessential amino acids, 1 mM sodium pyruvate, 15 mM HEPES solution, 100 U/mL penicillin and 100 μg/mL streptomycin as previously described51 (link)52 (link). The highly metastatic MDA-MB-231-HM.LNm5 cell line was generated through a two-step process whereby a cell population initially selected by 6 cycles of transplanting spontaneous MDA-MB-231 pulmonary metastases to the mammary fat pad44 (link) (designated MDA-MB-231-HM and kindly provided by ZM Shao and ZL Ou (Breast Cancer Institute, Fudan University, Shanghai, China) were inoculated into the mammary fat pad of a female BALB/c-SCID mouse, following which the axillary lymph node metastases were isolated. All experiments involving mice had approval from the Peter MacCallum Animal Experimentation Ethics Committee and were carried out in accordance with the Australian code for the care and use of animals for scientific purposes (8th edition, 2013). MDA-MB-231-HM.LNm5 cells were cultured in RPMI 1640 supplemented as above.
7
Quantitative Western Blot Protocol
8
Cell Line Maintenance and 1,6-Hexanediol Treatment
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MCF7 breast and A549 lung adenocarcinoma as well as DU-145 prostate cancer cell lines were purchased from ATCC and maintained in Roswell Park Memorial Institute 1640 (RPMI) medium (Biosera, Metro Manila, Philippines) complemented with 10% Fetal Bovine Serum (FBS) (EuroClone, Pero MI, Italy), 2 mM glutamine (Sigma-Aldrich, St. Louis, MO, USA), 0.01% streptomycin and 0.006% penicillin (Biowest, Nuaille, France). Cells were cultured under standard conditions in a 37 °C incubator at 5% CO2 and 95% humidity. When indicated, cells were treated with 1,6-hexanediol at a final concentration of 2% v/v for 30 min.
9
Cell Culture Conditions for HCT116 and MCF7
10
Isolation of Extracellular Vesicles from Cancer Cell Lines
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U-251 glioblastoma (GBM), MCF-7 breast, and A375 melanoma cancer cell lines were supplied by American Type Culture Collection (ATCC, Manassas, VA). Cell lines were cultured in their recommended culture medium32 containing 10% FBS and 1% penicillin–streptomycin at 37 °C in a 5% CO2 incubator. For isolation of EVs from CCM, U251, MCF7, and A375 cells were grown in T75 flasks to 90% cell confluence, followed by washing the cells twice with PBS. Culture medium with 10% EV-depleted FBS was added to cells for 24 h. CCM was centrifuged at 1, 000×g for 5 min at room temperature (RT) to discard cell debris before further processing. EV-depleted FBS was prepared by using the permeate of FBS filtered by tangential flow filtration (TFF) (MWCO: 300 kDa).
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