Between 250 and 1000 cells per well (optimized for each specific cell line) were plated in triplicate in 96 well plates in their respective FBS culture medium. At 24 h after plating, cells were changed to their corresponding culture medium supplemented with 5 or 10% CSS, and treated with either PX-12 (Sigma, M5324) or DMSO (Sigma, D2650) as the vehicle control. At 72 h following PX-12 treatment, luminescence was measured using the Cell TiterGlo Kit (Promega, Ref G7571), and read on a FilterMax F5 Microplate Reader (Molecular Devices, LLC). Data were normalized to luminescence values from vehicle-treated controls within each group (i.e., DMSO FBS or DMSO CSS), and plotted as % relative luminescence units (RLU). For oxidant treatments, cells were incubated with either H2O2 (Sigma, H1009) or methyl viologen dichloride hydrate (a.k.a paraquat, Sigma, 856177). For all in vitro experiments, cell culture dishes corresponding to a specific line were randomly assigned to the treatment groups.
Px 12
Manufactured by Merck Group
Sourced in United States
The PX-12 is a laboratory equipment designed for performing X-ray diffraction analysis. It is capable of analyzing the crystalline structure of materials by measuring the diffraction patterns of X-rays interacting with the sample. The PX-12 provides accurate and reliable data on the atomic arrangements and composition of various solid-state materials.
Automatically generated - may contain errors
Lab products found in correlation
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Celltiter glo kit, by Promega (1 mentions)
Methyl viologen dichloride hydrate, by Merck Group (1 mentions)
Filtermax f5 microplate reader, by Molecular Devices (1 mentions)
Hydrogen peroxide h2o2, by Merck Group (1 mentions)
N acetyl l cysteine, by Merck Group (1 mentions)
Celltiter 96 aqueous one cell proliferation assay, by Promega (1 mentions)
Sp600125, by Enzo Life Sciences (1 mentions)
Horseradish peroxidase conjugated secondary antibody, by Cell Signaling Technology (1 mentions)
Tunicamycin, by Merck Group (1 mentions)
Total eif2α, by Cell Signaling Technology (1 mentions)
E cadherin, by Cell Signaling Technology (1 mentions)
Tgf β, by R&D Systems (1 mentions)
Sirtinol, by Merck Group (1 mentions)
Angiotensin 2, by R&D Systems (1 mentions)
Hrp conjugated secondary antibody, by Santa Cruz Biotechnology (1 mentions)
Rhodamine conjugated secondary antibody, by Jackson ImmunoResearch (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
Gapdh, by Cell Signaling Technology (1 mentions)
Hdac4, by Abcam (1 mentions)
7 protocols using px 12
1
Cytotoxicity Evaluation of PX-12 and Oxidants
2
Oxidative Stress Modulation Protocols
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The AP-1 inhibitor (NDGA), NADPH oxidase inhibitor (DPI), Thioredoxin inhibitor (PX-12), and the antioxidants N-acetyl L-cysteine (NAC) and propyl gallate were obtained from Sigma. The JNK inhibitor (SP600125) was obtained from Enzo. NAC was initially reconstituted in DR 5% culture medium at 30 mM , its pH was adjusted to 7.0 using 1M NaOH solution and was filter-sterilised before use. Other inhibitors were reconstituted in DMSO (Sigma) or ethanol (propyl gallate) and appropriate solvent-only controls were included in all experiments. For some experiments, cells were treated with hydrogen peroxide (H2O2) (Sigma) diluted in culture medium at the indicated concentrations for 48 h. The effects of H2O2 on cell growth were quantified by measuring cell biomass using the CellTiter 96 AQueous One cell proliferation assay (Promega) and results were expressed as % cell growth of each condition in comparison to untreated controls, as detailed elsewhere.63 (link)
3
Molecular Mechanisms of Cellular Stress Response
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Tunicamycin, sirtinol, PX12, and fatty acids-bearing albumin (A4919, low endotoxin <0.1 ng/mg) were obtained from Sigma Chemical Company (St. Louis, MO, USA). TGF-β, angiotensin II and glucose were obtained from R&D Systems (Minneapolis, MN, USA). Antibody to GRP78 was acquired from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Antibodies to heme oxygenase-1, thioredoxin, total eIF2α (eukaryotic translation initiation factor-2α), phosphospecific-eIF2α (Ser51), α-smooth muscle actin, E-cadherin, and horseradish peroxidase-conjugated secondary antibody were purchased from Cell Signaling Technology (Beverly, MA, USA). Zinc protoporphyrin IX (Zn(II)PPIX) was obtained from Calbiochem (San Diego, CA, USA).
4
Mitochondrial Dynamics Regulation Protocol
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The phalloidin, JC-1 and DCFH-DA probes were bought from Life Technology (St. Louis, MO, USA). PX-12, DAPI and isoprenaline hydrochloride (Iso) were purchased from Sigma-Aldrich (USA). Rhodamine-conjugated secondary antibodies were obtained from Jackson ImmunoResearch Laboratories (West Grove, PA, USA). STVNa was obtained from Chemical Development Laboratories of Key Biological Pharmaceutical Company (Dongguan, China). Trizol was obtained from Generay (Shanghai, China). Antibodies against Drp1, Trx1, Prdx2 and GAPDH were acquired from Cell Signaling Technology (Beverly, MA, USA). HDAC4 was obtained from Abcam (Cambridge, MA, UK). The Fis1 antibody and HRP-conjugated secondary antibodies were acquired from Santa Cruz Biotechnology (Santa Cruz, CA, USA).
5
Characterization of Tellurium Compounds
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Biotin-maleimide, colchicine, DTNB (5,5′-Dithiobis(2-nitrobenzoic acid)), insulin, NADPH, paclitaxel (taxol), PDI, p-nitrophenyl phosphate and PX-12 were purchased from Sigma-Aldrich (Stockholm, Sweden). Recombinant HIV-1(IIIB) glycoprotein gp120 (CHO) was purchased from Immuno Diagnostics, Inc. (Woburn, MA) and Trx1,TrxR1 and the FkGRX-01 kit for Grx1 assay were from IMCO Corporation (Stockholm, Sweden). The compounds 6-(butyltelluro)-6-deoxy-β-cyclodextrin (TE-2), bis[4-(N,N-di(2-carbomethoxyethyl)amino)phenyl]telluride (TE-10), N,N-dimethyl-4-aminophenyl 3-phenoxypropyl telluride (TE-14) and 3-(butyltelluro)propanesulfonic acid sodium salt (TE-20) were synthesised as described previously31 (link)–33 (link) dissolved in dimethylsulfoxide (DMSO) at 25 mM and kept at −20 °C until use. The monoclonal anti-acetylated tubulin antibody (catalogue # T7451, clone 6-11B-1, Batch # 036M4856V 1.1 mg/ml) and anti-β-actin (catalogue # A5441, clone AC-15) were both from Sigma-Aldrich (St. Louis, MO). TZM-bl cells were obtained through the NIH AIDS Reagent Program, Division of AIDS, NIAID, NIH, from Dr. John C. Kappes, Dr. Xiaoyun Wu and Tranzyme Inc. and the persistently HIV-1 infected ACH-2 cells were obtained from Dr. Thomas Folks.
6
Preparation of Bioactive Compounds
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DTNB, NTZ, EGCG, rutin, bacitracin, and PX-12 were purchased from Sigma-Aldrich and dissolved in dimethyl sulfoxide (DMSO), ethanol, or water according to the manufacturer’s instructions.
7
Cytotoxicity and Apoptosis Induction in Colon Cancer Cells
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Chloroform, ethanol, methanol, span 60, Tween 60, cholesterol, dimethyl sulfoxide (DMSO), ultrapure sodium dodecyl sulfate (SDS), and Amicon® Ultra Centrifugal Filter Devices (Amicon Ultra-15 Membrane MWCO 30 KDa) were purchased from Merck company (Germany). Phosphate buffered saline tablet (PBS) provided by Bio Basic Company(Canada). The PX-12 (PubChem CID: 219,104), YC-1(PubChem CID: 5712), DEPC water, and dialysis membrane (MWCO 12 KDa) were obtained from Sigma Aldrich company(USA). Dulbecco's Modified Eagle's medium (DMEM), Trypsin–EDTA, and Penicillin–Streptomycin(pen-strep) were received fromBiosera company (France). Fetal bovine serum (FBS) was bought from Gibcocompany (USA)0.3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was purchased from Atocelcompany(Australia).ApoFlowEx FITC Kit (Annexin V/PI) was acquired from EXBIO company (Czech Republic). The flow cytometer used in this study is a FACScalibur belonging to BD Biosciencescompany (Canada). Sybr Green master mix real-time PCR was purchased from Ampliqoncompany (Denmark). Intestinal cancer cells (HT-29) and human foreskin fibroblast (HFF) were supplied from the National Center for Genetic and Biological Resources of Iran.
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