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    Isothermal Titration Calorimetry: A Biophysical Method to Characterize the Interaction between Label-free Biomolecules in Solution
    Author:  Andrea Saponaro, date: 08/05/2018, view: 154, Q&A: 0
    This protocol can be applied to analyze the direct interaction between a soluble protein and a target ligand molecule using Isothermal Titration Calorimetry (ITC, Malvern). ITC allows the biophysical characterization of binding between label-free, ...
    Inositol Phosphates Purification Using Titanium Dioxide Beads
    Authors:  Miranda SC Wilson and Adolfo Saiardi, date: 08/05/2018, view: 94, Q&A: 0
    Inositol phosphates (IPs) comprise a family of ubiquitous eukaryotic signaling molecules. They have been linked to the regulation of a pleiotropy of important cellular activities, but low abundance and detection difficulties have hampered our ...
    Determination of Storage (Starch/Glycogen) and Total Saccharides Content in Algae and Cyanobacteria by a Phenol-Sulfuric Acid Method
    Authors:  Tomáš Zavřel, Petra Očenášová, Maria A. Sinetova and Jan Červený, date: 08/05/2018, view: 80, Q&A: 0
    This is a protocol for quantitative determination of storage and total carbohydrates in algae and cyanobacteria. The protocol is simple, fast and sensitive and it requires only few standard chemicals. Great advantage of this protocol is that both ...
    Isothermal Titration Calorimetry: A Biophysical Method to Characterize the Interaction between Label-free Biomolecules in Solution
    Author:  Andrea Saponaro, date: 08/05/2018, view: 154, Q&A: 0
    [Abstract] This protocol can be applied to analyze the direct interaction between a soluble protein and a target ligand molecule using Isothermal Titration Calorimetry (ITC, Malvern). ITC allows the biophysical characterization of binding between label-free, non-immobilized and in-solution biomolecules by providing the stoichiometry of the interaction, the ...
    Inositol Phosphates Purification Using Titanium Dioxide Beads
    Authors:  Miranda SC Wilson and Adolfo Saiardi, date: 08/05/2018, view: 94, Q&A: 0
    [Abstract] Inositol phosphates (IPs) comprise a family of ubiquitous eukaryotic signaling molecules. They have been linked to the regulation of a pleiotropy of important cellular activities, but low abundance and detection difficulties have hampered our understanding. Here we present a method to purify and enrich IPs or other phosphate-rich metabolites from ...
    Determination of Storage (Starch/Glycogen) and Total Saccharides Content in Algae and Cyanobacteria by a Phenol-Sulfuric Acid Method
    Authors:  Tomáš Zavřel, Petra Očenášová, Maria A. Sinetova and Jan Červený, date: 08/05/2018, view: 80, Q&A: 0
    [Abstract] This is a protocol for quantitative determination of storage and total carbohydrates in algae and cyanobacteria. The protocol is simple, fast and sensitive and it requires only few standard chemicals. Great advantage of this protocol is that both storage and total saccharides can be determined in the cellular pellets that were already used for ...
    Measuring CD38 Hydrolase and Cyclase Activities: 1,N6-Ethenonicotinamide Adenine Dinucleotide (ε-NAD) and Nicotinamide Guanine Dinucleotide (NGD) Fluorescence-based Methods
    [Abstract] CD38 is a multifunctional enzyme involved in calcium signaling and Nicotinamide Adenine Dinucleotide (NAD+) metabolism. Through its major activity, the hydrolysis of NAD+, CD38 helps maintain the appropriate levels of this molecule for all NAD+-dependent metabolic processes to occur. Due to current advances and ...
    Two Different Methods of Quantification of Oxidized Nicotinamide Adenine Dinucleotide (NAD+) and Reduced Nicotinamide Adenine Dinucleotide (NADH) Intracellular Levels: Enzymatic Coupled Cycling Assay and Ultra-performance Liquid Chromatography (UPLC)-Mass Spectrometry
    [Abstract] Current studies on the age-related development of metabolic dysfunction and frailty are each day in more evidence. It is known, as aging progresses, nicotinamide adenine dinucleotide (NAD+) levels decrease in an expected physiological process. Recent studies have shown that a reduction in NAD+ is a key factor for the ...
    In vitro Enzymatic Assays of Histone Decrotonylation on Recombinant Histones
    Authors:  Rachel Fellows and Patrick Varga-Weisz, date: 07/20/2018, view: 308, Q&A: 0
    [Abstract] Class I histone deacetylases (HDACs) are efficient histone decrotonylases, broadening the enzymatic spectrum of these important (epi-)genome regulators and drug targets. Here, we describe an in vitro approach to assaying class I HDACs with different acyl-histone substrates, including crotonylated histones and expand this to examine the ...
    α-Synuclein Aggregation Monitored by Thioflavin T Fluorescence Assay
    Authors:  Michael M. Wördehoff and Wolfgang Hoyer, date: 07/20/2018, view: 335, Q&A: 0
    [Abstract] Studying the aggregation of amyloid proteins like α-synuclein in vitro is a convenient and popular tool to gain kinetic insights into aggregation as well as to study factors (e.g., aggregation inhibitors) that influence it. These aggregation assays typically make use of the fluorescence dye Thioflavin T as a sensitive ...
    BMV Propagation, Extraction and Purification Using Chromatographic Methods
    [Abstract] Brome mosaic virus (BMV) is a well-known plant virus representing single-stranded RNA (ssRNA) positive-sense viruses. It has been widely used as a model in multiple studies concerning plant virus biology, epidemiology and the application of viral capsids in nanotechnology. Herein, we describe a method for BMV purification based on ion-exchange and ...
    Assessment of Uptake and Biodistribution of Radiolabeled Cholesterol in Mice Using Gavaged Recombinant Triglyceride-rich Lipoprotein Particles (rTRL)
    Authors:  Anna Worthmann, Clara John and Joerg Heeren, date: 07/05/2018, view: 315, Q&A: 0
    [Abstract] The here described method can be used to estimate the uptake of orally provided cholesterol in mice. Briefly, mice are gavaged with radiolabeled cholesterol and 4 h later, organ distribution of the radiolabel is determined by liquid scintillation counting. The method has been applied successfully to determine dietary cholesterol handling of mice ...
    Fluorescent Labeling of Rat-tail Collagen for 3D Fluorescence Imaging
    Author:  Andrew D. D Doyle, date: 07/05/2018, view: 522, Q&A: 0
    [Abstract] Rat tail collagen solutions have been used as polymerizable in vitro three-dimensional (3D) extracellular matrix (ECM) gels for single and collective cell migration assays as well as spheroid formation. These 3D hydrogels are a relatively inexpensive, simple to use model system that can mimic the in vivo physical characteristics ...