Immunohistochemistry / edited by S. Renshaw.

Format:

Book

Contributor:

Renshaw, S. (Simon)

Series:

Methods express

Methods express series

Language:

English

Subjects (All):

Immunohistochemistry.

Immunohistochemistry--methods.

Medical Subjects:

Immunohistochemistry--methods.

Physical Description:

xxiv, 264 pages, 12 unnumbered pages of plates : illustrations (partly color) ; 25 cm.

Place of Publication:

Bloxham, Oxfordshire : Scion, 2007.

Summary:

Immunohistochemistry: Methods Express is structured to reflect the modern applications of immunohistochemistry, combining emphasis on the growing trend towards automation and high throughput technology, with clear descriptions of the more established methodology.

The book covers the key techniques, such as: Choice of antibodies and the selection of reporter labels, Immunostaining - specimen preparation, fixation, antigen retrieval, and immunohistochemical staining methods, Multiple immunostaining, Confocal microscopy, Electron microscopy, Image capture, analysis and quantification, Automation, Quality assurance and troubleshooting.

This book is a comprehensive research guide; every chapter discusses the merits and limitations of various approaches and then provides selected tried-and-tested protocols with useful hints and tips for success. Immunohistochemistry: Methods Express is an essential laboratory manual for researchers in all biomedical fields and at all levels, from postgraduate student to principal investigator.

Contents:

Color section xiii

Chapter 2 Antibodies for immunochemistry / Catherine Onley

1.1 Antibodies are a family of glycoproteins with a central role in the adaptive immune response 3

1.2 Isotypic, allotypic, and idiotypic variations contribute to the diverse repertoire of antibody structures 6

1.3 The structures of antibodies are optimized for antigen binding and effector functions 8

1.4 Multiple factors contribute to the antibody-antigen interaction, including antibody affinity, avidity, and specificity 10

2 Methods and approaches 11

2.1 The relationship between antibody affinity, avidity, and specificity 11

2.2 Antibody production 14

2.3 Recommended protocols 27

Chapter 3 The selection of reporter labels / Sarah Mardle

2 Methods and approaches 34

2.1 Enzymatic/chromogenic detection 34

2.2 Fluorescence detection 37

2.3 Blocking unwanted background signals 40

2.4 Chromogen development 40

2.5 Mounting media and slide storage 41

2.6 Storage of enzyme- and fluorochrome-conjugated reagents 41

2.7 Enzymatic or fluorescent? 41

Chapter 4 Immunochemical staining techniques / Simon Renshaw

1.1 Paraffin-embedded sections 45

1.2 Frozen tissue sections 46

1.3 Free-floating sections 46

1.4 Cytological specimens 47

1.5 Reproducible and accurate results 47

2 Methods and approaches 47

2.1 Specimen fixation 47

2.2 Processing 56

2.3 Specimen storage 58

2.4 Decalcification 58

2.5 Antigen retrieval 59

2.6 Counterstaining following immunochemical staining 61

2.7 Mounting following immunochemical staining 64

2.8 Slide storage following immunochemical staining 67

2.9 Tissue microarrays 67

2.10 Recommended protocols 68

3 Troubleshooting 95

Chapter 5 Multiple immunochemical staining / Ian William Jones, Adam Westmacott

1.1 Choosing an appropriate method 97

1.2 Experimental design 98

1.3 Appropriate controls 101

1.4 Multiple staining using same-species primary antibodies 102

2 Methods and approaches 107

2.1 Labels for light microscopy techniques 107

2.2 Labels for fluorescence and confocal microscopy techniques 109

2.3 Labels for electron microscopy techniques 109

2.4 Recommended protocols 110

3 Troubleshooting 124

Chapter 6 Confocal microscopy and immunochemistry / Matthew Cuttle

1.1 Do you need to use confocal microscopy? 128

1.2 How does the confocal microscope work? 129

2 Methods and approaches 131

2.1 Selecting fluorescent dyes 131

2.2 Setting up the light path on the microscope 132

2.3 Choosing the right lens 133

2.4 How to set the digital image settings 134

2.5 Getting ready to scan 135

2.6 Optimizing the image collection settings 136

2.7 Optimizing z stack image collection 139

2.8 The advantages of multiphoton confocal microscopy 140

2.9 Advanced techniques for imaging with multiple fluorochromes 141

2.10 Enhancing the final image 145

Chapter 7 Ultrastructural immunochemistry / Jeremy N. Skepper, Janet M. Powell

1.1 Fixation and its effect on antigen-antibody binding 150

1.2 Controls 153

1.3 Why do we need to use electron microscopy? 156

1.4 Quantification 156

2 Methods and approaches 157

2.1 Epoxy resin sections 158

2.2 The acrylic resins London Resin (LR) White and Gold 159

2.3 Freeze substitution and low-temperature embedding in Lowicryl HM20 162

2.4 Ultrathin thawed cryosections 163

2.5 Recommended protocols 166

Chapter 8 Image capture, analysis, and quantification / Jiahua Wu, Anthony Warford, David Tannahill

2 Methods and approaches 176

2.1 Image capture 176

2.2 Image analysis and quantification 183

2.3 Image data handling 198

Chapter 9 Quality assurance in immunochemistry / Peter Jackson

2 Methods and approaches 206

2.1 Fixation and tissue processing 206

2.2 Microtomy 209

2.3 Decalcification 211

2.4 Antigen retrieval 211

2.5 Immunochemical methodologies 216

2.6 Controls 218

2.7 Microscopic interpretation 221

2.8 Background staining 222

3 Troubleshooting 230

Chapter 10 Automated immunochemistry / Emanuel Schenck

1.1 Defining the needs 240

2 Methods and approaches 241

2.1 Overview of automated platforms for immunochemical staining 241

2.2 System contrasts 241

2.3 Other special features 246

2.4 System running costs 246

2.5 System failure safeguards 246.

Notes:

Includes bibliographical references and index.

ISBN:

1904842178

9781904842170

1904842038

9781904842033

OCLC:

61129823