AP39, a novel mitochondria-targeted H2S donor, was designed and synthesized by Medicilon Inc., as described previously [19 (link), 20 (link)]. Antimycin A, 0.25% trypsin, 2-deoxyglucose, oligomycin, carbonyl cyanide-4-(trifluoromethoxy)phenylhydrazone (FCCP), trichloroacetic acid, rotenone, 7-azido-4-methylcoumarin (AzMC), monobasic potassium phosphate, dibasic sodium succinate hexahydrate, adenosine 5′-triphosphate (ATP) disodium salt hydrate, fatty acid-free BSA, sodium hydrosulfide hydrate (NaSH·xH2O), N,N-diethyl-p-phenylenediamine sulfate, magnesium chloride, and iron(III) chloride (FeCl3) were purchased from Sigma-Aldrich Company. The monoclonal anti-Aβ antibody 6E10 was obtained from Covance Inc. (Princeton, NJ, USA). Drp1 was purchased from Novus Biologicals, Inc. Fis1 was purchased from Proteintech Group. Mfn1 and Mfn2 antibodies were purchased from Santa Cruz Biotech, and OPA-1 antibody was purchased from BD Transduction Laboratories. The secondary antibodies goat anti-mouse-HRP and donkey anti-rabbit-HRP antibodies were purchased from GE Healthcare. The Pierce BCA protein assay kit was purchased from Thermo Fisher Scientific (Waltham, MA, USA). Dulbecco's modified Eagle's medium (DMEM), 1% glutamine, and 1% penicillin were obtained from Invitrogen. All other chemicals were purchased from Amresco (Solon, OH, USA).
7 azido 4 methylcoumarin
Manufactured by Merck Group
Sourced in United States
7-Azido-4-Methylcoumarin is a chemical compound used as a fluorescent label in biological research. It is a coumarin derivative with an azido group attached to the 7-position. The compound exhibits fluorescent properties, making it useful for labeling and tracking applications in cell biology and biochemistry experiments.
Automatically generated - may contain errors
Lab products found in correlation
L cysteine, by Merck Group (3 mentions)
Oligomycin, by Merck Group (2 mentions)
2 deoxyglucose, by Merck Group (2 mentions)
Antimycin a, by Merck Group (2 mentions)
Rotenone, by Merck Group (2 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
Adenosine 5 triphosphate atp disodium salt hydrate, by Merck Group (1 mentions)
N n diethyl p phenylenediamine sulfate, by Merck Group (1 mentions)
Iron 3 chloride fecl3, by Merck Group (1 mentions)
Goat anti mouse hrp, by GE Healthcare (1 mentions)
Potassium phosphate monobasic, by Merck Group (1 mentions)
Donkey anti rabbit hrp, by GE Healthcare (1 mentions)
Magnesium chloride, by Merck Group (1 mentions)
Trypsin, by Merck Group (1 mentions)
Fatty acid free bsa, by Merck Group (1 mentions)
Dulbecco s modified eagle s medium dmem, by Thermo Fisher Scientific (1 mentions)
E coli dh5α, by Tiangen Biotech (1 mentions)
Lead nitrate, by Merck Group (1 mentions)
35 mm glass bottom dish, by Ibidi (1 mentions)
6 protocols using 7 azido 4 methylcoumarin
1
Mitochondrial Hydrogen Sulfide Donor Synthesis
2
Recombinant IscS Expression in E. coli
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The plasmid pET-28a (Novagen, United States) was used for IscS expression. E. coli DH5α (Tiangen, China) was used to amplify recombinant plasmids, and E. coli BL21(DE3) pLysS (Tiangen) was the host strain for IscS expression. E. coli BW25113 and the mutant strain BW25113 (ΔiscS) were kindly provided by Jingdan Liang from Shanghai Jiao Tong University (An et al., 2012 (link)). The mutant strain BW25113 (ΔcyuA) was kindly provided by Sheng Yang from Chinese academy of science. Sodium phenylpyruvate (PPNa) and diamide were obtained from TCI (Shanghai, China). Dithiothreitol (DTT), trans-4,5-dihydroxy-1,2-dithiane (DTToxi), 7-Azido-4-methylcoumarin, lead nitrate and L-cysteine were obtained from Sigma-Aldrich (St. Louis, United States). The other materials used in this study were purchased from Sangon (Shanghai, China). Unless otherwise specified, all chemicals were used as they were received.
3
Visualization of 6AF-Treated PNT2 Cells
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PNT2 cells were cultured in a 35 mm glass bottom dish (ibidi) until 20% confluency. Cells were treated with 200 μM of 6AF for 72 h at 37 °C in an atmosphere of 5% CO2. After 3 days, cells were fixed with 1% paraformaldehyde, “click” with a fluorescent tag 7-Azido-4-Methylcoumarin from Sigma-Aldrich with excitation at 387 nm and emission at 470 nm, and membrane stained with Cellmark dye (ThermoFisher, Waltham, MA, USA) with excitation at 522 nm and emission at 535 nm. Images were taken at VetMed Advance Imaging Facility under 100× magnification with oil submersion.
4
Biochemical Reagents for Research
5
Apoptosis Assays via Redox-Sensitive Compounds
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7-Azido-4-Methylcoumarin, L-cysteine, L-homocysteine, pyridoxal 5′-phosphate hydrate, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), propidium iodide (PI), and ribonuclease (RNase) were purchased from Sigma-Aldrich (St. Louis, MO, USA). The following antibodies were purchased from Cell Signaling Technology (Beverly, MA, USA): rabbit anti-Bax, anti-PARP, anti-Cyto C, and mouse anti-caspase 9. Mouse anti-MPST was purchased from Santa Cruz. Rabbit anti-CBS and anti-CTH were obtained from HUA BIO and ZEN BIO, respectively. Mouse anti-β-actin was obtained from Sigma-Aldrich. A protease inhibitor cocktail was purchased from Roche (Basel, Kanton Basel, Switzerland). Lipofectamine 2000 transfection reagent was ordered from Invitrogen (Carlsbad, CA, USA). A FITC annexin V apoptosis detection kit was purchased from BD Pharmingen (San Diego, CA, USA). The Caspase-Glo® 3/7 Assay and the CellTiter-Glo® Luminescent Cell Viability Assay were purchased from Promega (Madison, WI, USA).
6
Molecular Mechanisms of Fibrosis Inhibition
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Sodium hydrogen sulfide (NaHS), PPG (a CSE inhibitor) and 7-Azido-4-Methylcoumarin (AzMC) were purchased from Sigma-Aldrich (St. Louis, MO, USA). SKF38393 (a DR1 agonist) was obtained from Abcam (Cambridge, MA, USA). Y-27632 (a ROCK inhibitor) was obtained from MedChemExpress (Shanghai, China). The primary antibodies for anti-CSE, Cyclin D1, proliferating cell nuclear antigen (PCNA), p21 Cip/W AF -1 , collagen I (Col-1), collagen III (Col-3), matrix metalloproteinase 9 (MMP-9), osteopontin (OPN) and α-smooth muscle actin (α-SMA) were purchased from Proteintech (Wuhan, China). The anti-p-RhoA, t-RhoA, p-ROCK1, t-ROCK1 were from Affinity Biosciences (Cincinnati, OH, USA). The anti-DR1 antibody was from GeneTex (Irvine, CA, USA). Horseradish peroxidase-conjugated goat anti-rabbit IgG, goat anti-mouse IgG antibody, the Cell Counting Kit-8 (CCK-8) and anti-β-actin were obtained from Boster Bioengineering Limited Company (Wuhan, China). The EdU Cell Proliferation Assay Kit were obtained from Ribobio (Guangzhou, China). Fluo-4 AM were obtained from Beyotime Biotechnology (Shanghai, China). Enhanced ECL Chemiluminescent Substrate Kit was obtained from Yeasen Biotechnology (Shanghai, China). All other chemicals were from Solarbio (Beijing, China) or Beyotime Biotechnology.
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