Ethylenediaminetetraacetic acid was obtained from Fisher Scientific International, UK. Aged garlic extract was kindly provided by Wakunaga of America Company Ltd. Ribose, sodium dodecyl sulphate (SDS) and Tris (hydroxymethyl)-amino methane were obtained from BDH, UK. Acrylamide solution was obtained from Bio-Rad Laboratories (Hemel Hempstead, UK). Bromophenol blue was obtained from Serva, Germany. Ethanol, glacial acetic acid and mEthanol were obtained from Fisher Scientific International, UK. Dialysis tubing with a molecular weight cut off of 3.5 kDa and 67 kDa was obtained from Medical International Ltd Company (London, UK). Advanced glycation endproduct antibody (rabbit polyclonal to AGE antibody) and rabbit IgG secondary antibody (goat polyclonal to rabbit IgG) were purchased from Abcam, UK. All other reagents were of analytical reagent grade from Sigma-Aldrich Company (Poole, UK).
Acrylamide solution
Manufactured by Bio-Rad
Sourced in United States
Acrylamide solution is a liquid reagent used in various biochemical and molecular biology applications. It serves as a precursor for the formation of polyacrylamide gels, which are commonly used in techniques such as gel electrophoresis for the separation and analysis of biomolecules, including proteins and nucleic acids.
Automatically generated - may contain errors
Lab products found in correlation
Sulfo sanpah, by Thermo Fisher Scientific (4 mentions)
Ammonium persulfate, by Bio-Rad (3 mentions)
N n methylene bis acrylamide solutions, by Bio-Rad (3 mentions)
Acrylamide bis solution, by Bio-Rad (3 mentions)
Trypsin edta, by Thermo Fisher Scientific (3 mentions)
Bovine serum albumin (bsa), by Merck Group (2 mentions)
Glycine, by Bio-Rad (2 mentions)
Matlab, by MathWorks (2 mentions)
Ti e microscope, by Nikon (2 mentions)
Methanol, by Merck Group (2 mentions)
Sodium azide, by Merck Group (2 mentions)
Sodium dodecyl sulfate (sds), by Merck Group (2 mentions)
Sodium acetate, by Merck Group (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
T4 dna ligase, by Thermo Fisher Scientific (2 mentions)
Ethylenediaminetetraacetic acid, by Thermo Fisher Scientific (1 mentions)
Ethanol, by Thermo Fisher Scientific (1 mentions)
Methanol, by Thermo Fisher Scientific (1 mentions)
Glacial acetic acid, by Thermo Fisher Scientific (1 mentions)
Tetramethylethylenediamine, by Merck Group (1 mentions)
22 protocols using acrylamide solution
1
Characterization of Advanced Glycation Endproducts
2
Characterization of Food Additives
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Commercial food additive SiO2 (AEROSIL 200F, food grade) was purchased from Evonik Industries AG (Essen, Germany). Food grade titanium dioxide and zinc oxide was purchased from Tioxide Europe SRL. (Ternate, VA, Italy) and Spectrum Chemical Mfg. Corp. (New Brunswick, NJ, USA), respectively. Bovine serum albumin (BSA), casein sodium salt, maltose monohydrate, triton X-100, Bradford reagent, isopropyl-β-d-thiogalactopyranoside (IPTG), sodium dodecyl sulfate (SDS) and tetramethylethylenediamine (TEMED) were purchased from Sigma-Aldrich (St. Louis, MO, USA). BamHI and XbaI were purchased from New England Biolabs Inc. (Ipswich, MA, USA). Luria-Bertani (LB) broth and T4 lagase were obtained from BD Difco (Franklin Lakes, NJ, USA) and Promega Co (Madison, Wl, USA), respectively. Pfu polymerase and dNTP were purchased from Bioneer Co. (Daejeon, Korea). Waxy maize starch and sucrose were obtained from Yakuri Pure Chemicals (Kyoto, Japan) and Samyang Co. (Gyeonggi, Korea), respectively. Acrylamide solution (30%) was purchased from Bio-Rad Laboratory (Hercules, California, USA). Anodized aluminum oxide membranes (AnodiscTM, pore size 0.2 μm) and Ni-NTA agarose resin were purchased from Whatman (Maidstone, UK) and Qiagen Inc. (Valencia, CA, USA), respectively. Six processed foods were purchased from the local market.
3
Simvastatin Cytotoxicity Evaluation in Cell Culture
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Eagle’s Minimum Essential Medium (MEM), trypsin ethylendiaminetetraacetate, penicillin (10,000 U/mL), streptomycin (10 mg/mL), nonessential amino acid solution (100×), fetal calf serum (FCS), disposable culture flasks, Petri dishes (Corning), and filters (Millipore) were purchased from Euroclone (Milano, Italy). [6–3H]-thymidine, sodium salt (2 Ci/mmol) and molecular-weight protein standards were from Amersham. Isoton II was purchased from Instrumentation Laboratories (Milano, Italy). Sodium dodecyl sulfate (SDS), NNNN-tetra-methyl-ethylendiamine, ammonium persulfate, glycine, and acrylamide solution (30% T, 2.6%) were obtained from Bio-Rad Laboratories. Simvastatin in its lactone form (Merck, Sharp, & Dohme Research Laboratories, Rahway, NJ, USA) was dissolved in 0.1 M NaOH to give the active form, and the pH was adjusted to 7.4 by adding 0.1 M HCl. The solution was sterilized by filtration.
4
One-Dimensional SDS-PAGE Protein Separation
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The proteins in the supernatant samples were separated by one-dimensional (1D-) SDS-PAGE. In brief, 20 μL of supernatant samples were mixed with 5 μL SDS sample buffer (50 mM Tris-HCl (pH 6.8), 2% SDS, 10% glycerol, 0.1% bromphenol blue) and denatured at 90 °C for 10 min. The SDS-PAGE gel included a separating gel (10% “Acrylamide-Solution (30%)-Mix 37.5:1” (Bio-Rad, USA), 0.4 M Tris (pH 8.8), 0.1% SDS, 0.1% APS, 0.04% TEMED) and a stacking gel (4% “Acrylamide-Solution”, 0.125 M Tris (pH 6.8), 0.1% SDS, 0.05% APS, 0.1% TEMED). The protein separation according to their molecular weight was conducted at 150 V for one hour using a Protean II Cell system (BIO-RAD). After electrophoresis, the gel was stained with Coomassie Brilliant Blue R-250.
5
Polyacrylamide Polymerization on t-ZnO Tablets
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Example 3
Interconnected t-ZnO tablets were used as templates for polyacrylamide polymerization. A mixture of acrylamide solution (Bio-Rad, 40%, 1.00 mL), Bis solution (Bio-Rad, 2%, 10.0-250 μL), and ammonium persulfate solution (Sigma-Aldrich, 10%, aq., 30.0 μL) was filled up to a volume of 5.00 mL in a small beaker and degassed for 20 min in a desiccator. The solution was mixed with N,N,N′,N′-tetramethylethyldiamine (TEMED, Bio-Rad, 10.0 μL) and the calculated volume for complete coverage of each t-ZnO tablet was poured on the tablet. After 1 h of polymerization the substrate was washed with bidest. H2O.
6
Fabrication of Polyacrylamide Substrates
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Polyacrylamide substrates ranging from 0.08 kPa–119 kPa were fabricated as described previously [9] (link). Briefly, acrylamide solution (Bio-Rad) ranging from 3%–15% was mixed with N-N'- methylene-bis-acrylamide solutions (Bio-Rad) ranging from 0.05%–1.2% and then polymerized between a glutaraldehyde-activated glass surface and hydrophobic coverslip using 10% ammonium persulfate (Bio-Rad) and 1/2000 TEMED (Sigma-Aldrich). Polymerized substrates were then activated for protein conjugation with the water-soluble, heterobifunctional crosslinker Sulfo-SANPAH at 0.5 mg/mL (Pierce Chemical Co.) under UV exposure followed by functionalization with human plasma fibronectin (Millipore Corp.) at a nominal surface density of 2.6 µg/cm2.
7
Lactoferrin Conjugation and Characterization
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Glycyrrhizin (ammonium salt, purity 95%), sodium periodate (NaIO4), lactoferrin human (Lf), sodium carbonate (Na2CO3), sodium bicarbonate (NaHCO3), sodium cyanoborohydride (NaBH3CN), fluorescein isothiocyanate isomer 1 (FITC), 2-mercaptoethanol and tetramethylethylenediamine (TEMED), and water (HPLC grade) wer.e purchased from Sigma-Aldrich (St. Louis, MO, USA). 4′,6-diamidino-2-phenylindole (DAPI) was purchased from Vector Laboratories (Burlingame, CA, USA). NP40, cell extraction buffer, and Lysotracker® were purchased from Invitrogen (Waltham, MA, USA). Phenylmethylsulfonyl fluoride (PMSF), acetonitrile, and methanol (HPLC grade) were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Paraformaldehyde (4%) was purchased by Wako (Kanto, Saitama, Japan). Protease inhibitor and Calcein AM (Cat #: BMD00064) were purchased from Abbkine (Wuhan, China). Sodium dodecyl sulfate was purchased from Affymetrix (Waltham, MA, USA). Acrylamide solution (30%) and ammonium persulfate (APS) were purchased by BIO-RAD (Hercules, CA, USA). Coomassie blue solution was purchased from Abcam (Cambridge, UK).
8
Hydrogel Preparation and Cell Imaging
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Polyacrylamide hydrogels were produced as previously described52 (link). Briefly, acrylamide solution (Bio-Rad) was mixed with N-N'- methylene-bis-acrylamide solutions (BioRad). A glutaraldehyde-activated glass surface and hydrophobic coverslip was then used for polymerization. Polymerized substrates were then activated for protein conjugation with the heterobifunctional crosslinker Sulfo-SANPAH at 0.5 mg/mL (Pierce Chemical Co.) under UV exposure for 15 min. The substances were then washed with HEPES buffer, and then functionalized with fibronectin. Using atomic force microscopy, gels were determined to have elastic moduli of ~120 kPa. Functionalized gels were washed with PBS, and ES2 shSCRM or shDDR2 were plated. Live cells were imaged on a Nikon Ti-E microscope with a temperature and humidity controlled incubation chamber (LiveCell). Data were collected by 3×3 tiling of 10× regions of view. Time-lapse imaging was performed with 20-minute frame intervals for a 6 hour duration. Image analysis was performed using Matlab (The Mathworks, Natick MA) and Image J. Cell area was computed for each cell in each field of view using custom software to detect cell boundaries. 3 independent experiments conducted, and pooled results quantified (n>200 for each condition). Data was plotted using R graphing library36 .
9
Adipocyte Differentiation Assay
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DIM, dimethyl-sulfoxide (DMSO), insulin, dexamethasone, isobutyl-methylxanthine (IBMX) and RIPA buffer were purchased from Sigma-Aldrich (St. Luois, MO). Dulbecco’s modified Eagle’s medium-high glucose (DMEM-HG), phosphate buffered saline solution (PBS), fetal bovine serum (FBS), non-essential amino acids and antibiotic-antimycotic solutions from Gibco Life-technologies (Grand Island, NY, USA). Enzyme linked immunosorbent assay (ELISA) kits to measure MCP-1, IL-6, and TNF-α were obtained from R&D Systems (Minneapolis MN). Trizol reagent, Moloney Murine Leukemia Virus (M-MLV) reverse transcriptase, and random primers from Invitrogene (Grand Island, NY, USA). Taqman gene expression assays, and Taqman gene expression master mix were purchased from Life Technologies Applied Biosystems (Grand Island, NY, USA). Protease, and phosphatase inhibitors (Complete minitablets) were purchased from Roche Applied Science (Basel, Switzerland). Acrylamide solution, and polyvinylidene difluoride (PVDF) membranes from Bio-Rad (CA, USA). Primary antibodies anti-IRS-1 pY612 were obtained from Abcam Incorporation (Cambridge, UK). Primary antibodies: anti-Akt-1/PKB pT308, and anti-GAPDH; and secondary antibodies: HRP-conjugated anti-IRS-1 pY612 anti-mouse, anti-Akt-1/PKB pT308 anti-rabbit, and anti-GAPDH anti-mouse, from Santa Cruz Biotechnology (Dallas, Texas, USA).
10
Fabrication of Collagen-Coated Polyacrylamide Gels
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The PA-gels were synthesized and pretreated using methods reported elsewhere [46 (link)]. In brief, 40% acrylamide solution (Bio-Rad), 2% bis-acrylamide solution (Bio-Rad), and water were mixed in the ratio of 179:15:6 (0.6 kPa). Then, 1% v/v ammonium persulfate (APS, Thermo Fisher Scientific) and 0.1% v/v methylethylenediamine (TEMED, Thermo Fisher Scientific) were added into the mixture. An adequate amount of the mixed solution was added onto the chloro-silanated glass surface and then covered by amino-silanated coverslips. After solidification, the PA gels stuck to the coverslip were detached from the glass together for later use. Before assays, the PA gel was coated with 200 μg/mL collagen type I (Thermo Fisher Scientific) via the crosslinker sulfosuccinimidyl 6-(4′-azido-2′-nitrophenylamino) hexanoate (sulfo-SANPAH; Thermo Fisher Scientific).
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