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P2989 Sigma

Proteoglycan Protocols

  •  ISBN-10 0-89603-759-2

  •  ISBN-13 978-0-89603-759-5

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publication info   R.V. Iozzo, ed., Humana Press, 2001, 576 pp., hard cover
mfr. no.   Humana

Description

General description

This collection of step-by-step protocols enables researchers to isolate intact proteoglycans from tissues and cultured cells, to establish the composition of their carbohydrate moieties, and to generate strategies for prokaryotic and eukaryotic expression. There are also detailed techniques for the suppression of specific proteoglycan genes, for the detection of mutant cells and their degradation products, and for studying specific interactions between proteoglycans and extracellular matrix proteins and with growth factors and their receptors.

Table of Contents

I. Isolation and Purification. Isolation of Proteoglycans from Cell Cultures and Tissues, Isolation of Proteoglycans from Tendon, Purification of Proteoglycans from Mineralized Tissues, Purification of Perlecan from Endothelial Cells, Isolation and Characterization of Nervous Tissue Proteoglycans, . Analysis of Proteoglycans and Glycosaminoglycans from Drosophila, Cartilage and Smooth Muscle Cell Proteoglycans Detected by Affinity Blotting Using Biotinylated Hyaluronan, Preparation of Proteoglycans for N-Terminal and Internal Amino Acid Sequence Analysis, High–Specific-Activity 35S-Labeled Heparan Sulfate Prepared from Cultured Cells, Selective Detection of Sulfotransferase Isoforms by the Ligand Affinity-Conversion Approach, Particulate and Soluble Glycosaminoglycan-Synthesizing Enzymes: Preparation Assay and Use, Disaccharide Composition of Hyaluronan and Chondroitin/Dermatan Sulfate: Analysis with Fluorophore-Assisted Carbohydrate Electrophoresis, Integral Glycan Sequencing of Heparan Sulfate and Heparin Saccharides, . Analytical and Preparative Strong Anion-Exchange HPLC of Heparan Sulfate and Heparin Saccharides, . Proteoglycans Analyzed by Composite Gel Electrophoresis and Immunoblotting, Quantitation of Proteoglycans in Biological Fluids Using Alcian Blue, Cellulose Acetate Electrophoresis of Glycosaminoglycans, . Disaccharide Composition in Glycosaminoglycans/Proteoglycans Analyzed by Capillary Zone Electrophoresis, Intact and Oligomeric Glycosaminoglycans Analyzed with Capillary Electrophoresis, II. Expression, Detection, and Degradation. Recombinant Expression of Proteoglycans in Mammalian Cells: Utility and Advantages of the Vaccinia Virus/T7 Bacteriophage Hybrid Expression System, Purification of Recombinant Human Decorin and Its Subdomains, Prokaryotic Expression of Proteoglycans, Proteoglycan Gene Targeting in Somatic Cells, Constitutive and Inducible Antisense Expression, Cell-Mediated Transfer of Proteoglycan Genes, Morphological Evaluation of Proteoglycans in Cells and Tissues, Expression and Characterization of Engineered Proteoglycans, . Intracellular Localization of Engineered Proteoglycans, . Selection of Glycosaminoglycan-Deficient Mutants, Xyloside Priming of Glycosaminoglycan Biosynthesis and Inhibition of Proteoglycan Assembly, Inhibition of Heparan Sulfate Synthesis by Chlorate, Detection of Proteoglycan Core Proteins with Glycosaminoglycan Lyases and Antibodies, . Proteoglycan Core Proteins and Catabolic Fragments Present in Tissues and Fluids, Degradation of Heparan Sulfate by Nitrous Acid, . Degradation of Heparan Sulfate with Heparin Lyases, Degradation of Chondroitin Sulfate and Dermatan Sulfate with Chondroitin Lyases, . In Vitro Assay for Hyaluronan Synthase, . Hyaluronidase Activity and Hyaluronidase Inhibitors: Assay Using a Microtiter-Based System, . Detecting Hyaluronidase and Hyaluronidase Inhibitors: Hyaluronan-Substrate Gel and -Inverse Substrate Gel Techniques, III. Interactions. Affinity Coelectrophoresis of Proteoglycan–Protein Complexes, . Binding Constant Measurements for Inhibitors of Growth Factor Binding to Heparan Sulfate Proteoglycans, Interaction of Proteoglycans with Receptor Tyrosine Kinases, Measuring Single-Cell Cytosolic Ca2+ Concentration in Response to Proteoglycans, Serum Amyloid A Peptide Interactions with Glycosaminoglycans: Evaluation by Affinity Chromatography, Interactions of Lipoproteins with Proteoglycans, Hyaluronan and Hyaluronan-Binding Proteins: Probes for Specific Detection, Novel Confocal-FRAP Analysis of Carbohydrate–Protein Interactions Within the Extracellular Matrix, Regulatory Roles of Syndecans in Cell Adhesion and Invasion, Optical Biosensor Techniques to Analyze Protein–Polysaccharide Interactions, Phage Display Technology to Obtain Antiheparan Sulfate Antibodies, Tissue-Specific Binding by FGF and FGF Receptors to Endogenous Heparan Sulfates, Index.

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