Quantification of the Binding of Insulin-like Growth Factor-I (IGF-I) and IGF Binding Protein-3 (IGFBP-3) Using Surface Plasmon Resonance

.......................... ..........................................................................II DEDICATION.......................... .........................................................................III AUTHOR'S ACKNOWLEDGEMENTS ........................................................................IV TABLE OF CONTENTS................ ........................................................................VI LIST OF FIGURES........................ .....................................................................IX LIST OF TABLES......................... ......................................................................X CHAPTER 1 – INTRODUCTION 1 1.0 Overview of the Chapter.................................................................................................... 1 1.1 The Insulin-like Growth Factor Family........................................................................... 1 1.2 Interaction of IGF-I and IGFBP-3 .................................................................................. 2 1.3 Importance of pH .............................................................................................................. 4 1.4 Binding Assays .................................................................................................................. 4 1.5 Surface Plasmon Resonance............................................................................................. 6 1.6 Overview of the Thesis .................................................................................................... 11 CHAPTER 2 – LITERATURE REVIEW 13 2.0 Overview of the Chapter.................................................................................................. 13 2.1 Data Analysis ................................................................................................................... 13 2.2 Review of Previous SPR Studies Using Insulin-like Growth Factors and Insulin-like Growth Factor Binding Proteins .......................................................................................... 22 2.3 Summary .......................................................................................................................... 29 CHAPTER 3 – LEICA MICROSYSTEMS BIO-SPR 9000 31 3.0 Overview of the Chapter.................................................................................................. 31 3.1 Leica SPR System Overview............................................................................................ 31 3.2 Solutions used for SPR Experiments.............................................................................. 32 3.3 Activation of the Carboxymethylated Dextran Slide and Immobilization of IGFBP-3 32 3.4 Preliminary Experiments ................................................................................................ 35 3.5 Initial Kinetic Experiments ............................................................................................. 36