The PLA-CS fibrous scaffold was manufactured using an electrospinning system including an electrical power supply, syringe-pushed pump, Teflon TM tubing, a blunt 18-G needle, and a collector. The high-voltage power was connected to the catheter tip, and aluminum foil was used as the collector owing to its electrical drag. PLA, CS, and trifluoroacetate were purchased from Sigma (St. Louis, MO, USA) and used without any further purification; 0.36 g CS and 0.18 g PLA were dissolved in 10 mL trifluoroacetate. The PLA-CS solution was poured into a 5-mL syringe connected to an 18-G needle at a 10-cm tip-to-collector distance, with the flow rate set to 1.2 mL/min by a pushed pump, and a polymer jet occurring under 25 kV. The fibers were continuously deposited on the aluminum foil to form a 3D nanofiber scaffold. The wettability of the PLA-CS fibrous scaffold was analyzed with a contact angle meter (FACE, CA-D). Fourier transform infrared (FTIR) spectrometry (Horiba FT-720) of the PLA-CS nanofibers was guided in the mid-IR region between 800 and 4000 cm -1 at a 16scan process. Prior to spectroscopic measurements, all specimens were mixed with potassium bromide at a ratio of 5:95, and then all pellets were fixed in the sample holder.
Trifluoroacetate
Manufactured by Merck Group
Sourced in United States
Trifluoroacetate is a chemical compound used in various laboratory applications. It serves as a reagent for the synthesis and purification of organic compounds. The core function of trifluoroacetate is to facilitate chemical reactions and separation processes in research and development settings.
Automatically generated - may contain errors
Lab products found in correlation
Acetonitrile, by Merck Group (3 mentions)
Dithiothreitol (dtt), by Merck Group (2 mentions)
Iodoacetamide, by Merck Group (2 mentions)
Ft 720, by Horiba (1 mentions)
Pageruler plus prestained protein ladder, by Thermo Fisher Scientific (1 mentions)
Dextran sulfate sodium (dss), by MP Biomedicals (1 mentions)
Sodium dodecyl sulfate (sds), by Merck Group (1 mentions)
Formic acid, by Thermo Fisher Scientific (1 mentions)
Acetonitrile, by Thermo Fisher Scientific (1 mentions)
Ultrapure water, by Thermo Fisher Scientific (1 mentions)
Acetone, by Thermo Fisher Scientific (1 mentions)
Ammonium bicarbonate, by Merck Group (1 mentions)
Urea, by Merck Group (1 mentions)
Proteominer low abundance protein enrichment kit, by Bio-Rad (1 mentions)
Azoxymethane, by Merck Group (1 mentions)
Pancreatin, by Promega (1 mentions)
Triethylammonium bicarbonate buffer, by Merck Group (1 mentions)
Tmt mass tagging kits and reagents, by Thermo Fisher Scientific (1 mentions)
Sodium acetate, by Merck Group (1 mentions)
Deae 52 cellulose, by Merck Group (1 mentions)
7 protocols using trifluoroacetate
1
Electrospinning of PLA-CS Nanofiber Scaffold
2
Proteomic Analysis of Colorectal Cancer
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Azoxymethane (Sigma, United States), dextran sulfate sodium (36–50 kDa, MP Biomedicals, United States), phosphate-buffered saline (Hyclone, United States), PageRuler Plus Prestained Protein Ladder (ThermoFisher, United States), TMT® Mass Tagging Kits and Reagents (ThermoFisher, United States), dithiothreitol (Sigma, United States), iodoacetamide (Sigma, United States), sodium dodecyl sulfate (Sigma, United States), urea (Sigma, United States), pancreatin (Promega, China), ammonium bicarbonate (Sigma, United States), UltraPure water (ThermoFisher, United States), triethylammonium bicarbonate buffer (Sigma, United States), acetonitrile (ThermoFisher, United States), formic acid (ThermoFisher, United States), acetone (ThermoFisher, United States), ProteoMiner Low-Abundance Protein Enrichment Kit (Bio-Rad, United States), and trifluoroacetate (Sigma, United States) were used in this study.
3
Isolation and Characterization of Choerospondias axillaris Leaf Polysaccharides
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The Choerospondias axillaris leaves were collected from Guangxi Medicinal Botanical Garden (Guangxi, China) and identified by Prof. Yong Tan. Glucose, arabinose, Galactose, galacturonic acid, glucuronic acid, ribose, xylose, Mannose, Rhamnose, Rhamnose (Rha) and fucose were purchased from Borui Sugar Biotechnology Co., Ltd. (Yangzhou, China). DEAE-52 cellulose, Sephadex G-100, Sodium acetate and Trifluoroacetate were purchased from Sigma Chemical Co (Shanghai, China). 3-methyl-1-phenyl-2-pyrazolin-5-one (PMP), nitroblue tetrazolium (NBT), 1,1-diphenyl-2-picrylhydrazyl (DPPH), phenazine methosulphate (PMS), reduced nicotinamide adenine dinucleotide (NADH), ferrozine, [2,2′-azinobis-(3-ethyl-benzothiazolin-6-sulfonic acid)] diammonium salt (ABTS) and 2,4,6-tris(2-pyridyl)-s-triazine (TPTZ) were purchased from Aladdin Chemical Co (Shanghai, China). Hydrogen peroxide (30%) water solution, PBS buffer solution (PH 7.2–7.4), 2,2-Di(4-tert-octylphenyl)-1-picrylhydrazyl and free radical and absolute ethanol (99.5%) were purchased from Guangxi Nanning Chenze Experimental Technology Company (Nanning, China).
4
Mass Spectrometry Proteome Analysis
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RNF183 precipitates obtained as described in the previous section were used as the proteome analysis sample. Precipitate-bound beads were suspended in 25.5 μl of bicarbonate ammonium (21.25 mM, Wako Pure Chemical Industries). After adding 1.5 μl of dithiothreitol (12.5 mM, Thermo Scientific), the mixture was incubated at 95°C for 5 min. After cooling to room temperature, 3 μl of iodoacetamide (25 mM, Wako Pure Chemical Industries) were added, and the mixture was incubated at room temperature for 20 min. Next, 10 μl of 30 ng/μl trypsin (Promega Corporation) were added, and the mixture was incubated at 37°C for 3 h. Another 10 μl of trypsin were added, and the mixture was incubated overnight at 30°C. Finally, 2.5 μl of trifluoroacetate (Sigma-Aldrich) were added to terminate the reaction, and the sample was desalted using a C18 Spin Column (Thermo Fisher Scientific), followed by concentration for 2 h on a SpeedVac. Concentrates were analyzed on a TripleTOF 5600+ System with Eksigent nanoLC (AB SCIEX, Framingham, MA, USA). Proteins were identified using the ProteinPilot Software (AB SCIEX).
5
Dried-droplet MALDI-TOF MS Analysis
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MS samples were prepared according to the Shimadzu manual (dried-droplet crystallization method; Axima® Performance Start Guide). Briefly, the matrix solution was prepared by dissolving 5 mg of MLP into 0.5 mL of acetonitrile/0.1% trifluoroacetate (Sigma-Aldrich, St. Louis, MO, USA) 1:1 solution in a clean centrifuge tube, followed by centrifugation at 20,000× g for 5 min. The supernatant was carefully mixed by pipetting with an equi-volume of the purified protein (analyte).
For MS detection, 1 μL of the mixed sample was applied onto a sample plate (Shimadzu, Kyoto, Japan) and allowed to dry completely under ambient conditions. An AXIMA Performance mass spectrometer (Shimadzu, Kyoto, Japan) was used in this study. Respective MS data acquisition was performed under the following conditions: power, 90; profiles, 50; shots, 1000; linear mode; with pulsed extraction optimized at 66,430.09 Da, equivalent to the molecular mass of BSA (Sigma, St. Louis, MO, USA) used as the calibration standard [18 (link)].
For MS detection, 1 μL of the mixed sample was applied onto a sample plate (Shimadzu, Kyoto, Japan) and allowed to dry completely under ambient conditions. An AXIMA Performance mass spectrometer (Shimadzu, Kyoto, Japan) was used in this study. Respective MS data acquisition was performed under the following conditions: power, 90; profiles, 50; shots, 1000; linear mode; with pulsed extraction optimized at 66,430.09 Da, equivalent to the molecular mass of BSA (Sigma, St. Louis, MO, USA) used as the calibration standard [18 (link)].
6
Quantitative Proteome Analysis Workflow
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Protease inhibitor cocktail tablets were purchased from Roche (Mannheim, Germany). Trypsin (sequencing grade) was purchased from Promega Corporation (Madison, WI). Ultra-centrifugal filters (10-kDa cutoff) were purchased from Sartorius (Göttingen, Germany). The C18 (octadecyl) solid-phase extraction disk was purchased from 3M Company (St. Paul, MN). The iTRAQ reagent kit was purchased from AB Sciex Corporation (Foster City, CA). The bicinchoninic acid protein assay kit, dithiothreitol, iodoacetamide, acetonitrile, trifluoroacetate, and formic acid were all purchased from Sigma-Aldrich Corporation (St. Louis, MO). All aqueous solutions were prepared using Milli-Q (Millipore, Billerica, MA) water.
7
HPLC Analysis of Ketoprofen in Plasma
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HPLC analyses were performed using a Dionex Ultimate 3000 Pump equipped with a Dionex ASI-100 Automated Sample Injector and with a UV-VIS lamp. Ketoprofen was extracted from the plasma samples using a method described elsewhere 25 with minor modifications. Briefly, methanol was added in a 3:1 v/v ratio to the plasma and, after vortexing the mixtures, the samples were centrifuged at 15,000 g for 6 min and the supernatants were transferred into HPLC vials. The HPLC was run in isocratic mode using a method already described in literature with slight modifications 25 . The mobile phases constituted of (A): deionized water with 1 % v/v trifluoroacetate (Sigma Aldrich, St. Louis, USA) and (B): 100 % acetonitrile (Sigma Aldrich, St. Louis, USA). The ratio of the mobile phase A:B was equal to 45:55 v/v. Samples were run over a Kinetex 5.0 µm XB-C18 100 Å, 100 x 4.6 mm column (Phenomenex ApS, Nordic Region, Vaerløse, Denmark) at 22 °C. The injected volume was 40 µL with a flow rate of 1 mL/min and a total run time per sample of 10 min. The absorbance was measured at 258 nm.
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