Cells cultured for 48 h in 2D culture plates, or for 14 d in 3DPS, were gently washed once in cell medium and fixated for 1 h at room temperature in 2.5% glutaraldehyde (Product ref G7651-10ML, Sigma-Aldrich Sweden AB, Stockholm, Sweden) diluted in cell medium. Scaffolds were washed once in TBS (50 mM Trizma-HCl, Sigma-Aldrich, Stockholm, Sweden; 150 mM NaCl, Merck, Stockholm, Sweden; pH 7.5) with 0.1 mM CaCl2 (Merck, Stockholm, Sweden), fixated for 1 h at room temperature in 1% osmium tetroxide (Sigma-Aldrich, Stockholm, Sweden) in TBS (50 mM Trizma-HCl (Sigma-Aldrich, Stockholm, Sweden); 150 mM NaCl, (Merck, Stockholm, Sweden); pH 7.5) with 0.1 mM CaCl2 (Merck, Stockholm, Sweden), rinsed in deionized water, plunge-frozen in liquid propane (EMS-002 Rapid Immersion Freezer, Electron Microscopy Sciences) and freeze-dried overnight (VirTis Sentry 2.0 Benchtop Freeze Dryer, SP Scientific, Warminster, PA, USA). Samples were mounted on aluminum stubs by adhesive carbon tabs (Agar Scientific) and coated with a 10 nm Au/Pd conducting thin film by sputter-coater (PECS Mod 682, Gatan Inc., Pleasanton, CA, USA). Samples were imaged by Zeiss SUPRA 40VP scanning electron microscope operated in secondary electron mode at 3.0–4.1 kV acceleration voltage, 9–17 mm working distance and 100–250,000× magnification range.
Trizma hcl
Manufactured by Merck Group
Sourced in United States, Sweden
Trizma HCl is a buffer compound used in various laboratory applications. It is a salt of tris(hydroxymethyl)aminomethane (Tris) and hydrochloric acid (HCl), which provides a stable pH environment for biological and chemical experiments.
Automatically generated - may contain errors
Lab products found in correlation
Trizma base, by Merck Group (9 mentions)
Ethylene glycol tetraacetic acid (egta), by Merck Group (3 mentions)
Hepes, by Merck Group (3 mentions)
Acetylcholine iodide, by Merck Group (2 mentions)
Mono iodinated 3 125i iodotyrosyl54 αbgt, by GE Healthcare (2 mentions)
Pronase e, by Merck Group (2 mentions)
Milli q water, by Merck Group (2 mentions)
Citric acid, by Merck Group (2 mentions)
Acetonitrile, by Thermo Fisher Scientific (1 mentions)
Sodium chloride (nacl), by Avantor (1 mentions)
Sodium hydroxide pellets, by Avantor (1 mentions)
Decylubiquinone, by Merck Group (1 mentions)
Phenylarsine oxide pao, by Merck Group (1 mentions)
3 4 5 dimethylthiazol 2 yl 2 5 diphenyltetrazolium bromide mtt, by Merck Group (1 mentions)
Acetic acid, by Fujifilm (1 mentions)
Ammonium acetate, by Fujifilm (1 mentions)
Nitric acid, by Fujifilm (1 mentions)
Ammonia solution, by Fujifilm (1 mentions)
L cysteine, by Fujifilm (1 mentions)
Protease inhibitor cocktail, by Takara Bio (1 mentions)
19 protocols using trizma hcl
1
Characterization of 2D and 3D Cell Cultures
2
Nitric Oxide Metabolite Quantification
3
Biomolecular Reagents for Research
4
Protocols for Acetylcholine Receptor Studies
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Trizma-HCl, EGTA, HEPES, Pronase E, acetylcholine iodide, Cyt, and all chloride salts were purchased from Sigma (USA). Mono-iodinated (3-[125I]iodotyrosyl54)-αBgt (∼2000 Ci/mmol) was from GE Healthcare. nAChR-enriched membranes from the electric organs of T. californica ray were kindly provided by Prof. F. Hucho (Free University of Berlin, Germany), GH4C1 cells transfected with human α7 nAChR were a gift from Eli-Lilly (USA). The expressed acetylcholine binding proteins (AChBP) from L. stagnalis and Aplysia californica were kindly provided by Prof. T. Sixma (Netherlands Cancer Institute, Amsterdam, the Netherlands). Venom samples from vipers V. nikolskii and V. ursinii were obtained in 2009 by milking the snakes kept in captivity at Tula zoo (the Tula Exotarium). The permission was issued by the zoo director S.A.Ryabov. The venom of krait B. fasciatus was collected from snakes obtained from the authorized, licensed local establishment for snake breeding and venom production with permission of its owner Mr. Ha Văn Ti n (Vinh Sh n, Vinh Tu' ng, Vinh Phúc province, Vietnam). Cobra (N. kaouthia) venom was obtained as described earlier [17] (link). All efforts were undertaken to minimize suffering of the animals while venom samples were collected. Venoms collected were dried over anhydrous CaCl2 and stored at −20°C until use.
5
Arsenic Toxicity Evaluation Protocol
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All reagents were of analytical grade. Milli-Q water (Millipore, Bedford, MA, USA) was used throughout the experiment. Trizma® HCl and Trizma® Base, phenazene methosulfate, decylubiquinone, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), and phenylarsine oxide (PAO) were purchased from Sigma (St. Louis, MO, USA). Sodium arsenite (iAsIII), dimethylarsinic acid [(CH3)2AsO(OH)] (DMAV) were purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan). Monomethylarsonic acid (MMAV) was obtained from Tri Chemicals (Yamanashi, Japan). The arsenic standard solutions were stored in the dark at 4 °C. Diluted standard solutions for experiments were prepared daily prior to use.
6
Analytical Preparation of Arsenic Compounds
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All reagents were of analytical grade. Milli-Q water (Millipore) was used throughout the experiment. Trizma® HCl and Trizma® Base were purchased from Sigma (St. Louis, MO, USA). Nitric acid, ammonium acetate, acetic acid, 28% ammonia solution, L-cysteine, sodium arsenite (iAsIII), sodium arsenate (iAsV), and dimethylarsinic acid [(CH3)2AsO(OH)] (DMAV) were purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan). Monomethylarsonic acid (MMAV) was obtained from Tri Chemicals (Yamanashi, Japan). The arsenic standard solution (1, 000 μg/mL) for ICP-MS was purchased from SPEX CentiPrep (Metuchen, NJ, USA). Stock solutions of all arsenic compounds (10 mmol/L) were prepared from the respective standard compounds. All stock solutions were stored in the dark at 4°C. Diluted standard solutions for analysis were prepared daily prior to use.
7
Fluorescent Particle-Based Toxin B Assay
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Fluorescent microparticles (500 nm), 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) were purchased from Thermo Fisher Scientific (Waltham, MA). Polystyrene carboxylate magnetic particles (292 nm) were purchased from Ademtech (Pessac, France). Magnets for capturing magnetic particles were from Dexter Magnetic Technologies (Elk Grove, IL). Microtiter plates (96 well clear bottom, half area black plate) were from Greiner Bio-One (Monroe, NC). Native Toxin B standard purified from C. difficile (ribotype 087) was purchased from List Laboratories (Campbell, CA). Mouse monoclonal antibodies raised against C. difficile toxin B were from BBI Solutions (Cardiff, UK) and Fitzgerald (Acton, MA). Bovine serum albumin (BSA), Casein, Trizma® base, Trizma®-HCl, Triton X-100 and OptiPrep were from Sigma-Aldrich (St. Louis, MO). Direct Black 19 was from Orient Corporation (Cranford, NJ). Protease inhibitor cocktail was from Takara Bio (Mountain View, CA).
8
Enzymatic Activity Assay Protocol
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Trizma base, trizma HCl, aminopeptidase N (APN), L-leupine-p-nitroanilide (L-p-NA), cAMP-specific PDE V (PDE5), diallyl trisulfide (DATS), diallyl disulfide (DADS), kaempferol (KAE), quercetin (QUE), and p-nitrophenyl phenyl phosphonate (p-NPPPh) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Methiin (MT) was purchased from Abcam (Cambridge, MA, USA). Isoalliin (IA) was purchased from DiagnoCine (Hackensack, NJ, USA). All other reagents used in this study were obtained commercially and were of high-quality grade.
9
Sperm Aneuploidy Assessment by FISH
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Ejaculated specimens were prepared for FISH aneuploidy assessment as previously described [32 (link)]. Five microliters of semen samples was smeared on glass slides and air-dried overnight. Slides were fixed in Carnoy’s fixative (3:1 methanol:acetic acid) at room temperature and subsequently placed in a 37 °C slide moat overnight. Sperm nuclei decondensation was carried out by incubating slides in 5 mmol/L dithiothreitol (DTT; Sigma Chemical Co., St. Louis, MO) in 0.1 M tris(hydroxymethyl)aminomethane (Trizma HCl; Sigma Chemical Co.), and then by 3 M sodium chloride and 300 mM tri-sodium citrate dehydrate (2× standard saline citrate; Vysis, Downers Grove, IL), pH 7.0. The decondensed slides were hybridized with fluorescent probes specific for chromosomes X, Y, 13, 15, 16, 17, 18, 21, and 22. Sperm nuclei were then counterstained with 4′,6-diamino-2-phenylindole (DAPI; Millipore, Temecula, CA, USA). A minimum of 1000 spermatozoa per specimen were analyzed under an Olympus BX61 fluorescent microscope at 1000×. Incidences of disomy, nullisomy, and diploidy were recorded using a computerized system (Applied Imaging, CytoVysion v3.93.2), with a <1.6% normal threshold [32 (link)].
10
Measuring Lipid Peroxidation via MDA
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MDA production was measured following the method previously described by Esterbauer and Cheeseman [22 ] to assess lipid peroxidation. The peritoneal fluid samples were diluted in Tris HCl buffer (TRIZMA HCl, Sigma Aldrich, São Paulo, Brazil) in distilled water (20 mM, pH 7.4), following homogenization and centrifugation at 10,000 rpm for 10 min at 4°C. To each sample, chromogenic reagent (10.3 mM 1-methyl-2-phenylindole in 3:1 acetonitrile) and HCl (37%) were added. Subsequently, they were incubated for 40 min at 45°C and centrifuged at 10,000 rpm for 5 min at 4°C. Absorbance was recorded at 586 nm using Spectrocopical UV/VIS analysis (Biotek, São Paulo, Brazil) and interpolated in a standard curve with 1,1,3,3-tetraethoxypropane. The results are shown in nmol/μl.
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