A Laboratory Guide to Biotin-Labeling in Biomolecule Analysis

T. Meier

ISBN 10: 3034873514 / ISBN 13: 9783034873512

Published by Springer, Basel, Birkhäuser Basel, Birkhäuser Mrz 2012, 2012

About this Item

This item is printed on demand - it takes 3-4 days longer - Neuware -2.1 Introduction . 32 2.2 Technical Procedures . 32 Chemical Synthesis . 35 2.3 Results and Discussion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 38 . . . . . . . . . . . . . Chemical Synthesis . 38 Binding to anti-CCK antisera and CCK-B receptors . 40 Binding to streptavidin agarose and photoelution . 40 Affinity chromatography . 41 2.4 Troubleshooting. 43 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44 . . . . . . . . . . . . . . . . . Purification of the Receptor for Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) using Biotinylated Ligands . 45 Summary . 45 3.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46 . . . . . . . . . . . . . . . . . 3.2 Technical Procedures . 48 Preparation of a Biotinylated Ligand . 48 Preparation of Membrane Fraction . 50 Solubilization of the PACAP Receptor . 51 Partial Purification of the PACAP Receptor . 52 Affinity Purification of the PACAP Receptor . 54 Final Purification of the PACAP Receptor . 56 Receptor-Binding Assay . 57 3.3 Results and Discussion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58 . . . . . . . . . . . . . 3.4 Troubleshooting . 61 Impurity in PACAP27-Cys-NH. 61 2 Low specific activity of the PACAP receptor despite a single band . 62 Acknowledgments . 62 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63 . . . . . . . . . . . . . . . . . Photoreactive Biotinylated Peptide Ligands for Affinity Labeling . ' 65 Summary . 65 4.1 Introduction. . . . . . . . . . 66 . 4.2 Technical Procedures . ' 67 Synthesis of a trifunctional photoactivatable biotinylating reagent . 67 Synthesis of photo reactive biotinylated peptide hormones . 67 Site-specific incorporation of biotin and photo labels in separate steps . 69 Photoaffinity labeling . 72 4.3 Results and Discussion . 76 Synthesis of photoactivatable insulins with permanent biotin labels . 76 Applications: Insulin . 78 VI Examples for other applications . 78 4.4 Troubleshooting. 79 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81 . . . . . . . . . . . . . . . . . . . 240 pp. Englisch.

Seller Inventory # 9783034873512

Bibliographic Details

• Title: A Laboratory Guide to Biotin-Labeling in Biomolecule Analysis
• Publisher: Springer, Basel, Birkhäuser Basel, Birkhäuser Mrz 2012
• Publication Date: 2012
• Language: English
• ISBN 10: 3034873514
• ISBN 13: 9783034873512
• Binding: Taschenbuch
• Condition: Neu

About this title

Synopsis

2.1 Introduction ...32 2.2 Technical Procedures ...32 Chemical Synthesis ...35 2.3 Results and Discussion...38 ...Chemical Synthesis ...38 Binding to anti-CCK antisera and CCK-B receptors ...40 Binding to streptavidin agarose and photoelution ...40 Affinity chromatography ...41 2.4 Troubleshooting...43 References ...44 ...Purification of the Receptor for Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) using Biotinylated Ligands ...45 Summary ...45 3.1 Introduction ...46 ...3.2 Technical Procedures ...48 Preparation of a Biotinylated Ligand ...48 Preparation of Membrane Fraction ...50 Solubilization of the PACAP Receptor ...51 Partial Purification of the PACAP Receptor ...52 Affinity Purification of the PACAP Receptor ...54 Final Purification of the PACAP Receptor ...56 Receptor-Binding Assay ...57 3.3 Results and Discussion...58 ...3.4 Troubleshooting ...61 Impurity in PACAP27-Cys-NH...61 2 Low specific activity of the PACAP receptor despite a single band ...62 Acknowledgments ...62 References ...63 ...Photoreactive Biotinylated Peptide Ligands for Affinity Labeling ..." 65 Summary ...65 4.1 Introduction...66 . 4.2 Technical Procedures ..." 67 Synthesis of a trifunctional photoactivatable biotinylating reagent ...67 Synthesis of photo reactive biotinylated peptide hormones ...67 Site-specific incorporation of biotin and photo labels in separate steps ...69 Photoaffinity labeling ...72 4.3 Results and Discussion ...76 Synthesis of photoactivatable insulins with permanent biotin labels ...76 Applications: Insulin ...78 VI Examples for other applications ...78 4.4 Troubleshooting...79 References ...81 ...