Table of Contents

Table of Contents

TABLE OF CONTENTS………………………………………………………................I

ABSTRACT……………………………………………………………………………….III

LIST OF TABLES….…………………………………………………………...............IV

LIST OF FIGURES…….……………………………………………………..................V

NOMENCLATURE…………………………………………………………..................VI

1.         INTRODUCTION………………………………………………………. ...1

1.1      CHARACTERISTICS OF THE PRION DISEASES……………1

1.1 .1        General Features of the Prion Diseases………………..1

1.1.2         Prion Diseases: Etiology, Transmission, Clinical                                Presentation and Treatment……………………………...2

                                  

1.2      MOLECULAR PROPERTIES OF THE PRION PROTEIN……5

1.2.1         Structure of the Prion Protein…………………………….5

1.2.2         Function of the Prion Protein…………………………. …7

1.2.3         Genetics of the Prion Protein…………………………….8

1.2.4         Prions in Yeast and Fungi………………………………..9

1.3      CELLULAR TRAFFICKING OF THE PRION PROTEIN ……10

1.4      THE PRINCIPLES AND APPLICATIONS OF THE ATOMIC 

            FORCE MICROSCOPY ………………………………………..13

      

1.4.1           Atomic Force Microscope……………………………….13

1.4.2           General Principles of AFM………………………………13

1.5      AMYLOID IN PRION DISEASES………………………………15

                 1.5.1          Formation of Amyloid……………………………………15

     1.5.2          Amyloid under AFM……………………………………..17

                      

           

 2.     MATERIALS AND METHODS……………………………………….....19

2.1      MATERIALS……………………………………………………...19

2.1.1         Chemicals, Buffers, Solutions, Reagents and Media...19

2.1.2            Disposable Materials…………………………………… 19

2.1.3            Antibodies…………………………………………………20

2.1.4            Cell Lines………………………………………………….20

2.1.5            Hardware………………………………………………….20

2.1.6            Software…………………………………………………..20

                                  

2.2      METHODS………………………………………………………..21  

2.2.1         Cell Fixation for AFM and Immunofluorescence……...21

2.2.2         Immunofluorescence Detection of PrP^c / PrP^Sc ………21 

2.2.3            Imaging of Immunofluorescence Labeled Samples….22

2.2.4            Atomic Force Microscopy Setup……………………….22

2.2.5            Image Processing………………………………………..22

3.       RESULTS…………………………………………………………………23

                        3.1      INDIRECT IMMUNOFLUORESCENCE OF PrP ON THE                                         SURFACE OF NORMAL AND INFECTED N₂A CELLS…….23

                        3.2      CELL SURFACE STRUCTURES IN SCRAPIE-INFECTED                                                AND NON-INFECTED CELL CULTURES……………………24

4.       DISCUSSION…………………………………………………………….35

REFERENCES…………………………………………………………………………..38

HONOUR STATEMENT……………………………………………………................43