PtoMBD purified with affinity chromatography was buffer exchanged with 20 mM sodium phosphate, pH 7.4, containing 500 mM NaCl and concentrated using a Vivaspin Turbo 4 (10,000 MWCO) ultracentrifugation unit (Merck Millipore). The concentrated enzyme solution was loaded onto a HiLoad 16/600 Superdex 200 prep grade gel-filtration column (GE Healthcare) and eluted with the same buffer at a flow rate of 0.6 ml/min using an ÄKTA pure 25 chromatography system (GE Healthcare). The elution of protein was monitored by UV absorption at 280 nm. A standard curve for calibration was obtained using a gel filtration markers kit (Sigma–Aldrich) containing blue dextran (2000 kDa), thyroglobulin (669 kDa), apoferritin (443 kDa), β-amylase (200 kDa), alcohol dehydrogenase (150 kDa), bovine serum albumin (66 kDa), and carbonic anhydrase (29 kDa) and potassium ferricyanide (Wako).
Potassium ferricyanide
Manufactured by Fujifilm
Sourced in Japan
Potassium ferricyanide is a chemical compound used in various photographic and laboratory applications. It serves as a key component in the development of black-and-white photographic prints and film. Potassium ferricyanide is also utilized in various analytical and diagnostic testing procedures within laboratory settings.
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Lab products found in correlation
Potassium ferrocyanide, by Fujifilm (4 mentions)
Blue dextran, by Merck Group (1 mentions)
Kta pure 25 chromatography system, by GE Healthcare (1 mentions)
Gel filtration markers kit, by Merck Group (1 mentions)
Bovine serum albumin (bsa), by Fujifilm (1 mentions)
Triton x 100, by Merck Group (1 mentions)
Basal medium eagle bme, by Merck Group (1 mentions)
Trypsin solution, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Paraformaldehyde (pfa), by Merck Group (1 mentions)
Dimethyl sulfoxide (dmso), by Fujifilm (1 mentions)
Anti inos, by Santa Cruz Biotechnology (1 mentions)
Anti cox 2, by Santa Cruz Biotechnology (1 mentions)
Anti β actin, by Santa Cruz Biotechnology (1 mentions)
Malvidin, by Extrasynthese (1 mentions)
2 amino 2 hydroxymethyl 1 3 propanediol tris, by Fujifilm (1 mentions)
Methyl thiazolyl tetrazolium (mtt), by Fujifilm (1 mentions)
Nadph, by Merck Group (1 mentions)
Hypoxanthine, by Merck Group (1 mentions)
Xanthine oxidase, by Merck Group (1 mentions)
8 protocols using potassium ferricyanide
1
Purification and Characterization of PtoMBD Enzyme
2
Biochemical Analysis of Anti-inflammatory Effects
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Malvidin was purchased from Extrasynthese Co. (Genay, France). Basal medium eagle (BME), Triton X-100, and paraformaldehyde were obtained from Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). H2O2, calcium and magnesium-free PBS, MTT, dimethyl sulfoxide (DMSO), dimethyl formaldehyde, potassium ferrocyanide, Nonidet P-40 (NP-40), potassium ferricyanide, 2-amino-2-hydroxymethyl-1,3-propanediol (Tris), and phenylmethane sulfonyl fluoride (PMSF) were purchased from Wako Pure Chemical Industries Ltd. (Osaka, Japan). FBS and trypsin solution were obtained from Life Technologies Inc. (Grand Island, NY, USA), and 5-bromo-4-chloro-3-indoyl-b-D-galactoside was obtained from Nakalai (Kyoto, Japan). Anti-iNOS, anti-COX-2, and anti-β-actin were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA), and goat anti-rabbit IgG horseradish peroxidase-conjugated secondary antibodies were obtained from Amershan Corp. (Arlington Heights, IL, USA).
3
Antioxidant Assay Protocol
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GSH, Cys, NAC, HCS, Met, and potassium ferricyanide were purchased from Wako Chemical (Tokyo, Japan). TEMPOL, SOD from human erythrocytes, hypoxanthine, xanthine oxidase, NADH, and NADPH were purchased from Sigma-Aldrich (St. Louis, MO). Other chemicals used were of analytical grade. As basic solvents for the reaction mixtures, 100 mM phosphate buffer (PB) containing 0.05 mM DTPA adjusted to pH 7.4 was prepared. Deionized water (deionization by the Milli-Q system) was used to prepare PB.
4
Senescence-Associated β-Galactosidase Assay
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hiHepPCs in monolayer cultures were fixed with PBS containing 2% formaldehyde (Nacalai Tesque) and 0.2% glutaraldehyde (Nacalai Tesque), and senescence-associated β-galactosidase activities in hiHepPCs were analyzed as described previously45 (link). Briefly, the fixed cells were washed in PBS and incubated for 16 h at 37 °C with a mixture of 0.5 mg mL–1 X-gal (Anatrace) and 40 mM citric acid/sodium phosphate buffer (pH 6.0) containing 5 mM potassium ferricyanide (Wako), 5 mM potassium ferrocyanide (Wako), and 2 mM magnesium chloride (Wako).
5
Enzymatic Biofuel Cell Construction
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GDH (EC 1.1.1.47) was purchased from Toyobo (Osaka, Japan), Di (EC 1.8.1.4), and bilirubin oxidase (EC 1.3.3.5) from Myrothecium sp. were purchased from Amano Enzyme (Nagoya, Japan) and NAD and AQ2S were purchased from Oriental Yeast (Tokyo, Japan) and Tokyo Chemical Industry (Tokyo, Japan), respectively. PLL used for construction of the anode (MW > 300,000), glutaraldehyde and 1-pyrenebutyric acid N-hydroxysuccinimide ester (NHS-pyrene) were all purchased from Sigma Aldrich (St. Louis, MO, USA). PLL used for the cathode (Mw approximately 8000) was purchased from Peptide Institute Inc., Ltd. (Osaka, Japan). Glucose, potassium ferricyanide and phosphate buffer were purchased from Wako (Osaka, Japan). All other chemicals were of reagent grade quality.
6
Fabrication of Lateral Flow Immunoassay Strips
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Acrylic plates were purchased from Mitsubishi Rayon Co., Ltd. (Tokyo, Japan). Platinum (50 μm diameter) and silver (500 μm diameter) wires were purchased from Nilaco Co., Ltd. (Tokyo, Japan). Nitrocellulose membrane (Hi-Flow TM Plus, HF180), nitrocellulose membrane with the polyester backing (Hi-Flow TM Plus, HF240), sample pad and absorbent pad (Sure Wick ® ) were purchased from Millipore Co., Ltd. (Bedford, MA).
Monoclonal mouse anti-human albumin antibody (albumin antibody), polyclonal anti-human albumin antibody labeled with glucose oxidase (GOx labeled antibody) and albumin were purchased from Sigma. The enzymes used for measuring creatinine were creatininase (256 U mg -1 , Toyobo Co., Ltd., Tokyo, Japan), creatinase (13 U mg -1 , Toyobo), and sarcosine oxidase (16.5 U mg -1 , Toyobo). Casein, potassium ferrocyanide and potassium ferricyanide were obtained from Wako Pure Chemicals (Osaka, Japan). All other chemicals used were analytical reagents or laboratory grade.
Monoclonal mouse anti-human albumin antibody (albumin antibody), polyclonal anti-human albumin antibody labeled with glucose oxidase (GOx labeled antibody) and albumin were purchased from Sigma. The enzymes used for measuring creatinine were creatininase (256 U mg -1 , Toyobo Co., Ltd., Tokyo, Japan), creatinase (13 U mg -1 , Toyobo), and sarcosine oxidase (16.5 U mg -1 , Toyobo). Casein, potassium ferrocyanide and potassium ferricyanide were obtained from Wako Pure Chemicals (Osaka, Japan). All other chemicals used were analytical reagents or laboratory grade.
7
Cysteamine-Bound Polystyrene Particle Manipulation
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Chemicals Cysteamine (Sigma-Aldrich), potassium ferrocyanide (Wako), potassium ferricyanide (Wako), N-hydroxysuccinimide (NHS, Sigma-Aldrich), N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDAC, Sigma-Aldrich), silver nitrate (Wako), and sodium citrate (Sigma-Aldrich) were all reagent grade and used as purchased. Carboxylated polystyrene (PS) particles with 10 μm diameter dispersed in water (Polyscience, polybead ® carboxylate microspheres) were used as particles which were bound by Cysteamine and pulled by the electromagnetophoretic force. A silica capillary with 100 μm inner diameter (GL Science) was used as an optical capillary cell. 3-Aminopropyltrimethoxysilane (Sigma-Aldrich) was used for amination of the capillary wall. Other chemicals were all of reagent grade. Water was purified by a Milli-Q system (Merck Millipore).
8
Analytical-grade Reagents for Electrochemical Experiments
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All the chemicals were analytical grade and were used as received. Hexaammineruthenium(III) chloride was purchased from Sigma-Aldrich. Potassium ferricyanide was obtained from Wako Pure Chemical Industries, Ltd. (Japan).
KH2PO4, Na2HPO4 and sulfuric acid were purchased from Kanto Chemical. We prepared 100 mM phosphate buffer, which contained 100 mM KH2PO4 and Na2HPO4. We prepared a fresh concentration of H2O2 with 0.2 to 28 mM phosphate buffer (pH 7.0). Ultrapure water was used in all the experiments.
KH2PO4, Na2HPO4 and sulfuric acid were purchased from Kanto Chemical. We prepared 100 mM phosphate buffer, which contained 100 mM KH2PO4 and Na2HPO4. We prepared a fresh concentration of H2O2 with 0.2 to 28 mM phosphate buffer (pH 7.0). Ultrapure water was used in all the experiments.
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