The human breast carcinoma cell lines T47D and MCF7 were obtained from the American Type Culture Collection (Rockville, MD, USA). T47D and MCF7 cells were routinely grown in RPMI 1640 supplemented with l -glutamine (2 mM) and 10% foetal bovine serum. All-trans-RA was obtained from Sigma Chemical Co. (St. Louis, MO, USA). Retinoic acid stock solution was dissolved in dimethyl sulfoxide (DMSO) at a concentration of 10−2 M and maintained at −20°C, protected from light and in an inert atmosphere. The synthetic agonist retinoids, selective for RARα (BMS753), RARβ (BMS453) and RARγ (BMS961), and synthetic antagonist retinoids selective for RARα (BMS195614; Tocris Bioscience, Bristol, UK) were kindly provided by Dr. Hinrich Gronemeyer (IGBMC, Illkirch, France). Agonist and antagonist retinoids were diluted in ethanol and added to the culture medium to give a final concentration of 10−6 M. In control cultures, the DMSO or ethanol vehicle was added at the same final dilution. All experiments with retinoids were performed in reduced room light.
Bms961
Manufactured by Bio-Techne
Sourced in Spain, United Kingdom
BMS961 is a high-performance liquid chromatography (HPLC) system designed for analytical and preparative applications. It features a modular design, allowing for customization to meet specific research needs. The system includes a solvent delivery module, an autosampler, a column compartment, and a variety of compatible detectors. BMS961 is capable of handling a wide range of sample types and can be used for the separation, identification, and quantification of a variety of analytes.
Automatically generated - may contain errors
Lab products found in correlation
Am580, by Bio-Techne (3 mentions)
Bms195614, by Bio-Techne (2 mentions)
Bms753, by Bio-Techne (2 mentions)
Vp 16, by Merck Group (2 mentions)
Er50891, by Bio-Techne (2 mentions)
Cd2665, by Bio-Techne (2 mentions)
Bms 453, by Bio-Techne (1 mentions)
Mcf 7, by American Type Culture Collection (1 mentions)
Sulforhodamine, by Merck Group (1 mentions)
Mm11253, by Bio-Techne (1 mentions)
Cd3254, by Bio-Techne (1 mentions)
Bms493, by Bio-Techne (1 mentions)
Uvi3003, by Bio-Techne (1 mentions)
Bms614, by Bio-Techne (1 mentions)
Bms948, by Bristol-Myers Squibb (1 mentions)
Le135, by Bio-Techne (1 mentions)
7 protocols using bms961
1
Breast Cancer Cell Line Characterization
2
Generating DAPT-resistant Cell Lines
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A list of the characteristics, origin and growth conditions of the cell lines used in the study is available in Supplementary Methods . To obtain DAPT-resistant cell lines, MB-157 cells were cultured in the presence of the γ-secretase inhibitor (1 μM) for 40 days. Surviving cells were diluted in medium and replated at low density to isolate single-cell derived colonies. These cell cultures were grown in medium containing DAPT (1 μM) for 83 days. Two growing clones were isolated to obtain an equivalent number of DAPT-resistant cell lines (MB-157RCL7A and MB-157RCL15A). MB-157 cells stably over-expressing RARβ-targeting shRNAs were obtained by lentiviral infection of constructs based on the pGreenPuro-shRNA system (SBI, System Bioscences). The sequence of the RARβ-targeting shRNAs is available in Supplementary Methods . Lentiviral infected cells were subjected to puromycin (0.5 µg/mL) selection for the isolation of the shRARβ expressing cells.
The sensitivity of cell lines to the anti-proliferative action of ATRA was evaluated with the ATRA-score index [17 (link),18 (link)]. The following chemicals were used: ATRA (Sigma-Aldrich), AM580 (Tocris), BMS961 (Tocris), UVI2003 (a kind gift of Dr. Angel De Lera, Universidade de Vigo, Spain), ER50891 (Tocris), DAPT (Sigma), PF-03084014 (Pfeizer) and VP16 (SIGMA).
The sensitivity of cell lines to the anti-proliferative action of ATRA was evaluated with the ATRA-score index [17 (link),18 (link)]. The following chemicals were used: ATRA (Sigma-Aldrich), AM580 (Tocris), BMS961 (Tocris), UVI2003 (a kind gift of Dr. Angel De Lera, Universidade de Vigo, Spain), ER50891 (Tocris), DAPT (Sigma), PF-03084014 (Pfeizer) and VP16 (SIGMA).
3
Retinoid Receptor Modulation in Breast Cancer
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The following compounds were used: ATRA (Sigma-Aldrich, https://www.sigmaaldrich.com ), AM580 (Tocris, http://www.tocris.com ), BMS961 (Tocris), ER50891 (Tocris), CD2665 (Tocris), and UVI2003 (a kind gift of Dr. Angel De Lera, Universidade de Vigo, Spain). Sulforhodamine was from Sigma-Aldrich Co. A list of the cell lines, their characteristics, and origin is available in Supplementary Table S1 . The plasmid constructs used for RARα3 over-expression in MDA-MB453 cells and knockdown in SKBR3 cells are described below.
4
Preparation and Application of Beta-1,3-Glucan Beads
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Beta-1,3-glucan beads were prepared as previously described [27 (link)]. atRA was purchased from Sigma-Aldrich (Germany) and dissolved in absolute ethanol. The RARα-agonist BMS753, the RARγ-agonist BMS961, as well as the RARα antagonist BMS195614 and the RARγ antagonist MM11253 were purchased from Tocris Bioscience (UK). Monoclonal mouse anti-human Dectin-1 MAB1859 (clone #259931) antibody was purchased from R&D Systems (Germany). Mouse IgG2B isotype control antibody was purchased from eBioscience (UK). APC-conjugated polyclonal goat anti-mouse antibody and APC-conjugated monoclonal mouse anti-human CD14 antibody were purchased from BD Biosciences (Germany). Polyclonal rabbit anti-Galectin-3 SC-20157 antibody was purchased from Santa Cruz (USA) and polyclonal rabbit anti-Actin (20–33) antibody was purchased from Sigma-Aldrich (Germany). HRP conjugated goat anti-rabbit IgG (H+L) antibody was purchased from Dianova (Germany).
5
Receptor-mediated Transcriptional Regulation
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RXR is referred to RXRα hereafter. The pSG5-based Gal-RARα LBD (Gal-RARα), Gal-RXR LBD (Gal-RXR), Gal-TIF2 NRID (Gal-TIF2), Gal-SMRT NRID (Gal-SMRT), Gal-NCoR NRID (Gal-NCoR), RARα LBD-VP16 (RARα-VP16), RXRΔAB-VP16 (RXR-VP16), RXRΔAB, RARαΔAB, RXR, and RARγ expression vectors, and the (17m)5x-βGlob-Luc and the (RARE)3x-tk-Luc reporter genes have been described [52 (link),53 (link),54 (link)]. CD3254, UVI3003, LG100754 (LG754), LE135, CD2665, BMS961, BMS614, and BMS493 were from Tocris. BMS948 was provided by Bristol-Myers Squibb (New York, NY, USA) and AGN192870 (AGN870) by Galderma (Lausanne, Switzerland). UVI3002 was a gift of Angel de Lera (University of Vigo, Vigo, Spain). Am580 and TTNPB were from Sigma France (Saint Quentin Fallavier, France).
6
FACS Purification of Germ Cells
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Germ cell populations were purified from testes of Aldh1a1-3ser-/-, Rara;b;gSpg–/– and Rxra;b;gSpg–/– mice by FACS and characterized as described previously [13 (link),22 ]. Organotypic cultures of testes from Aldh1a1-3ser-/- mice were also as described previously, except that the RARG-selective agonist BMS961 at 10–7M (Tocris Bioscience) was used to activate RAR signaling instead of BMS753 [13 (link)].
7
Establishing DAPT-resistant Cell Lines
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A list of the characteristics, origin and growth conditions of the cell lines used in the study is available in Supplementary Methods. To obtain DAPT-resistant cell-lines, MB-157 cells were cultured in the presence of the γ-secretase inhibitor (1 µM) for 40 days. Surviving cells were diluted in medium and replated at low density to isolate single-cell derived colonies. These cell cultures were grown in medium containing DAPT (1 µM) for 83 days. Two growing clones were isolated to obtain an equivalent number of DAPTresistant cell-lines (MB-157RCL7A and MB-157RCL15A). HCC-1599 and MB-157 cells stably overexpressing RARβ-targeting shRNAs were obtained by lentiviral infection of constructs based on the pGreenPuro-shRNA system (SBI, System Bioscences). The sequence of the RARβ-targeting shRNAs is available in Supplementary Methods. Lentiviral infected cells were subjected to puromycin (0.5 µg/ml) selection for the isolation of the shRARβ expressing cells. The sensitivity of cell-lines to the antiproliferative action of ATRA was evaluated with the ATRA-score index (see Supplementary Methods) (17) .
The following chemicals were used: ATRA (Sigma-Aldrich), AM580 (Tocris), BMS961 (Tocris), UVI2003 (a kind gift of Dr. Angel De Lera, Universidade de Vigo, Spain), LE135 (Tocris), DAPT (Sigma), PF-03084014 (Pfeizer) and VP16 (SIGMA).
The following chemicals were used: ATRA (Sigma-Aldrich), AM580 (Tocris), BMS961 (Tocris), UVI2003 (a kind gift of Dr. Angel De Lera, Universidade de Vigo, Spain), LE135 (Tocris), DAPT (Sigma), PF-03084014 (Pfeizer) and VP16 (SIGMA).
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