NMR data were obtained with a Bruker AMX-600 (Germany). HRESIMS was performed on a Thermo LTQ Orbitrap-Discovery (United States). Acid hydrolysis results were analyzed by Agilent GC 7890A/5975C (United States). UV was recorded on a Hitachi U-2910 (Japan). IR were acquired using a Perkin Elmer Spectrum 400 (United States). Preparative HPLC used an Agilent 1260 instrument equipped with a Cosmosil ODS C18 column (5 μm, 10 mm × 250 mm). Optical rotations were measured on a Rodolph Autopol Ⅰ Polarimeter (United States); column chromatographies were performed with 80–100 and 200–300 mesh silica gel (Qingdao Haiyang) and Sephadex LH-20 (Pharmacia, United States). TLC was carried out on GF254 plates (Qingdao Haiyang). Human MDA-MB-231 cells were purchased from ATCC (VA, United States). D-(+)-glucose and tert-butylhydroquinone (tBHQ) were bought from Sigma Chemical (MO, United States).
Human mda mb 231 cells
Manufactured by American Type Culture Collection
Sourced in United States
Human MDA-MB-231 cells are a well-characterized adherent cell line derived from a breast adenocarcinoma. These cells are commonly used for in vitro research applications.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
D glucose, by Merck Group (1 mentions)
Spectrum 400, by PerkinElmer (1 mentions)
U 2910, by Hitachi (1 mentions)
Ltq orbitrap discovery, by Thermo Fisher Scientific (1 mentions)
Amx 600, by Bruker (1 mentions)
Tert butylhydroquinone tbhq, by Merck Group (1 mentions)
Mcf 10a, by American Type Culture Collection (1 mentions)
Dmem f12, by American Type Culture Collection (1 mentions)
Mda mb 231, by American Type Culture Collection (1 mentions)
Tc20 automated cell counter, by Bio-Rad (1 mentions)
Trypan blue, by Thermo Fisher Scientific (1 mentions)
Dmem culture medium, by Thermo Fisher Scientific (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
Glutamine, by Thermo Fisher Scientific (1 mentions)
Normocin, by Thermo Fisher Scientific (1 mentions)
Streptomycin p s, by Thermo Fisher Scientific (1 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions)
Raw264.7 cells, by Thermo Fisher Scientific (1 mentions)
Ifn γ, by Merck Group (1 mentions)
5 protocols using human mda mb 231 cells
1
Isolation and Characterization of Bioactive Compounds
2
Establishing Breast Cell Lines for Research
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Cell lines and culturing MDA-MB-231 (malignant) and MCF 10A (non-malignant) breast cell lines were purchased from ATCC and maintained in DMEM (Dulbecco's Modi ed Eagle Medium) and DMEM-F12 respectively. DMEM was supplemented with 10% fetal bovine serum (FBS) and DMEM-F12 was supplemented with 5% horse serum (HS). In experiments using SEVs, culture media were supplemented with SEV depleted serum (SEV -). To prepare FBS SEV -and HS EV -, the sera were ultracentrifuged at 100,000 x g overnight, and the supernatant was collected. Cells were cultured at 37 º C in 5 % of CO 2 atmosphere. Cell number was counted using a TC20 automated cell counter (Bio-Rad, Hercules, CA, USA) with 0.4% trypan blue (Thermo Scienti c, Waltham, MA, USA) prior experiments in order to check cell viability. pLenti-GFP-CD63 plasmid, previously described by Hoshino et. al. [56] , was used to make MDA-MB-231 cells stably expressing GFP-CD63. Human MDA-MB-231 cells (ATCC) and 293 FT packaging cells were grown in DMEM + 10% FBS. 293 FT cells transfection, viral harvest, and transduction of MDA-MB-231 cells were performed as previously described [57] . Transduced cells were selected with 4 µg/ml puromycin for 7 days.
3
Culturing MDA-MB-231 Cancer Cells
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The MDA-MB-231 cell line is well established as a model for cancer research related to gene regulation. The MDA-MB-231 cell line is also being used for lipid research. Human MDA-MB-231 cells were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA), and then grown in DMEM culture medium (Gibco, Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% fetal bovine serum (Gibco, Thermo Fisher Scientific, Waltham, MA, USA), 2 mM glutamine (Gibco, Thermo Fisher Scientific, Waltham, MA, USA), 1 mM sodium pyruvate, 10 mM HEPES, 100 μg/mL Normocin, 50 U/mL penicillin and 50 μg/mL streptomycin (P/S; Gibco, Thermo Fisher Scientific, Waltham, MA, USA) and incubated at 37 °C (with humidity) in 5% CO2.
4
Induction and Polarization of Macrophages for Cancer Research
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Human acute monocytic leukemia cells, THP-1 cells, mouse macrophages, RAW 264.7 cells, human MDA-MB-231 cells, and mouse 4T1 cells were obtained from American Type Culture Collection (ATCC, Manassas, VA, USA). THP-1 cells, RAW 264.7 cells, human MDA-MB-231 cells, and mouse 4T1 cells were cultured in RPMI-1640 medium (Gibco, Waltham, MA, USA) containing 10% fetal bovine serum (Gibco, Waltham, MA, USA) at 37 °C in an incubator of 5% CO2 and 95% air. To induce THP-1 cells differentiate into macrophages, THP-1 cells were incubated with 100 ng/ml phorbol 12-myristate 13-acetate (PMA; Sigma-Aldrich) for 48 h. To obtain ferroptosis-induced breast cancer cells, MDA-MB-231 cells and 4T1 cells were cultured with ferroptosis inducer, erastin (10 µM) for 24 h.
PMA-induced THP-1 cells and RAW 264.7 cells were cultured with five ng/mL lipopolysaccharide (Sigma) and 100 U/mL IFN- γ (Sigma) for 24 h for M1 polarization. To obtain M2 macrophages, PMA-induced THP-1 cells and RAW 264.7 cells were cultured with 10 ng/mL IL-4 (Sigma) for 24 h.
PMA-induced THP-1 cells and RAW 264.7 cells were cultured with five ng/mL lipopolysaccharide (Sigma) and 100 U/mL IFN- γ (Sigma) for 24 h for M1 polarization. To obtain M2 macrophages, PMA-induced THP-1 cells and RAW 264.7 cells were cultured with 10 ng/mL IL-4 (Sigma) for 24 h.
5
Human Breast Cancer Cell Culture
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The Human MDA-MB-231 cells and MCF-7 cells were obtained from the American Type Culture Collection (ATCC, Rockville, MD, USA) and cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% heated-activated fetal bovine serum (FBS), 100 U/mL penicillin, and 100 μg/mL streptomycin. The cells were subcultured every three days and maintained in a humidified incubator at 37 °C with a 5% CO2 atmosphere.
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