Patel / Arya / Shergill Basic Science Techniques in Clinical Practice

1. Auflage 2008
ISBN: 978-1-84628-740-4
Verlag: Springer
Format: PDF
Kopierschutz: 1 - PDF Watermark

E-Book, Englisch, 150 Seiten

A complete guide to implementing research projects for anyone in the medical professions. This book covers all the main areas, allowing anyone to set up and complete research projects. The techniques outlined here can easily be adapted to clinical projects. Written by international authors to provide a flavor from many institutions, the book's appeal is cross-sectional, both at hospital and primary care levels worldwide. Providing cutting-edge information in an accessible manner, and containing diagrams and easy-to-follow step-by-step guides, this is the first guide of its kind. It contains a complete section on setting up and funding research projects.

Autoren/Hrsg.

Arya, M.

Patel, Hitendra R. H.

Arya, Manit

Shergill, Iqbal

Inhaltsverzeichnis

1;Foreword;5
2;Preface;7
3;Contents;9
4;Contributors;11
5;Research Governance;15
5.1;INTRODUCTION;15
5.2;WHY WE NEED RESEARCH GOVERNANCE;16
5.3;HISTORY OF DEVELOPMENT OF RESEARCH GOVERNANCE1,2;16
5.4;SALIENT FEATURES OF THE RESEARCH GOVERNANCE FRAMEWORK;19
5.5;CONCLUSION;20
5.6;References;20
6;Designing Health Studies;22
6.1;ESSENTIALS OF A STUDY DESIGN Specifying the Research Question;22
6.2;Selection of Subjects;22
6.3;Specifying the Primary Outcome;22
6.4;Inclusion of a Control Group;23
6.5;Confounding;23
6.6;Sample Size;24
6.7;Bias;24
6.8;Writing a Protocol;25
6.9;TWO TYPES OF STUDIES;25
6.10;Randomized Controlled Trials;25
6.11;Types of Trials;26
6.12;Observational Studies;27
6.13;References;30
7;Immunohistochemistry;32
7.1;INTRODUCTION;32
7.2;BASIC IMMUNOHISTOCHEMISTRY;32
7.3;METHODOLOGY;34
7.4;Fixation;35
7.5;Antigen Retrieval;35
7.6;Antibodies;36
7.7;Staining Methods/Detection Systems;37
7.8;Enzyme Labels and Chromogens;40
7.9;Animal Tissue;41
7.10;TROUBLESHOOTING/OPTIMIZATION Optimizing Primary Antibodies;41
7.11;Endogenous Enzyme Activity;42
7.12;Endogenous Biotin;43
7.13;Washing;43
7.14;References;43
8;Cell Culturing: A Beginner’s Guide to Understanding the Basics of Cell Culturing;45
8.1;INTRODUCTION;45
8.2;Primary Cells;45
8.3;Cell Lines;45
8.4;FUNDAMENTALS OF CELL CULTURING;46
8.5;What Happens When an Infection Occurs?;46
8.6;Media: Their Function in Cell Culturing;47
8.7;Support Cells;48
8.8;Incubation;49
8.9;CELL EXTRACTION;49
8.10;Handling Tissue Biopsies;49
8.11;Fibroblast Extraction;49
8.12;CONCLUSION;51
8.13;References;51
9;Flow Cytometry;52
9.1;WHAT IS FLOW CYTOMETRY?;52
9.2;HISTORICAL ASPECTS OF FLOW CYTOMETRY AND THE FLOW CYTOMETER;53
9.3;FLUORESCENCE;53
9.4;LIGHT SCATTER;55
9.5;COMPONENTS OF THE MODERN FLOW CYTOMETER Fluidic System;55
9.6;Optical System and Analysis;55
9.7;Color Assignment;56
9.8;DATA ANALYSIS Histograms and Dot Plots;57
9.9;Gating and Negative Controls;58
9.10;PRINCIPLES OF FLOW CYTOMETRY SAMPLE PREPARATION;60
9.11;References;60
10;Western, Northern, and Southern Blotting;62
10.1;INTRODUCTION: THE HISTORY OF SOUTHERN, NORTHERN, AND WESTERN BLOTTING;62
10.2;BASIC PRINCIPLES AND METHODS: ELECTROPHORESIS, GEL BLOTTING, AND DETECTION;63
10.3;Gel Electrophoresis: Size Separation;63
10.4;Sample Preparation: Southern and Northern;63
10.5;Sample Preparation: Western;63
10.6;Agarose Electrophoresis: Southern and Northern;64
10.7;Sodium Sodecylsulfate–Polyacrylamide Gel Electrophoresis: Western;65
10.8;Blotting: Transfer to Solid Support;66
10.9;Detection and Localization: Complementarity and Hybridization;66
10.10;Blocking;68
10.11;Hybridization;68
10.12;Washing;68
10.13;Detection;68
10.14;Stripping;70
10.15;KEY POINTS;70
10.16;References;70
11;Fluorescent In Situ Hybridization;72
11.1;INTRODUCTION;72
11.2;BASIC PRINCIPLES;72
11.3;METHODOLOGICAL ASPECTS OF FISH Probe and Sample Preparation;73
11.4;DNA UNMASKING: PRETREATMENT AND DIGESTION De- waxing and Rehydration;74
11.5;Acid Permeabilization;74
11.6;Reducing Agents;74
11.7;Protease Digestion;74
11.8;Probe and Target DNA Denaturation;75
11.9;Probe and Target DNA Hybridization;75
11.10;POSTHYBRIDIZATION WASH AND VISUALIZATION Posthybridization Wash;76
11.11;Visualization and Scoring/Interpretation of Results;76
11.12;QUALITY ASSURANCE;76
11.13;Slide Pretreatment;76
11.14;Acid Permeabilization;77
11.15;Pretreatment (Reducing Agent);77
11.16;Protease Digestion;77
11.17;Denaturation;78
11.18;Hybridization;78
11.19;Posthybridization Wash;78
11.20;Visualization and Scoring Slides;79
11.21;References;79
12;Quantitative Reverse Transcriptase Polymerase Chain Reaction;80
12.1;INTRODUCTION;80
12.2;UNDERSTANDING THE FUNDAMENTALS;81
12.3;DISCUSSION OF METHODS TO QUANTIFY REAL-TIME POLYMERASE CHAIN REACTION RESULTS;84
12.4;CONTROLS IN QUANTITATIVE REVERSE TRANSCRIPTASE POLYMERASE CHAIN REACTION: USE OF THE HOUSEKEEPING” GENE;86
12.5;AMPLICON DETECTION STRATEGIES;86
12.6;DESIGNING A PRIMER, PROBE, AND AMPLICON;90
12.7;PUSHING THE TECHNOLOGY FURTHER: MULTIPLEX QUANTITATIVE REVERSE TRANSCRIPTASE POLYMERASE CHAIN REACTION;91
12.8;EQUIPMENT REVIEW;91
12.9;CONCLUSION;93
12.10;References;94
13;Proteonomics: High-Throughput Structural Biology — Methods for Cloning, Protein Expression, and Purification;100
13.1;INTRODUCTION;100
13.2;CURRENT APPROACHES HTP Cloning;101
13.3;Expression Systems;103
13.4;Expression Conditions;104
13.5;Expression and Solubility Screening;105
13.6;Puri.cation;105
13.7;ADDITIONAL APPROACHES;106
13.8;References;109
14;DNA and Tissue Microarrays;112
14.1;DNA MICROARRAY Methodology;112
14.2;Data Analysis;114
14.3;Sample Preparation Issues;115
14.4;Clinical Applications;115
14.5;TISSUE MICROARRAY Construction;116
14.6;Advantages and Criticisms of Tissue Microarrays;117
14.7;Applications;118
14.8;Quantitative Analysis;119
14.9;DNA and tissue microarray: key points;120
14.10;References;120
15;Basic Scientific Techniques in Recording Cellular Data;123
15.1;ELECTRICAL RECORDING SETUP;123
15.2;EXTRACELLULAR RECORDINGS;124
15.3;INTRACELLULAR RECORDINGS;125
15.4;MORPHOLOGY OF SINGLE CELLS BY INTRACELLULAR STAINING;126
15.5;OPTICAL RECORDINGS;127
15.6;CALCIUM-SENSITIVE OPTICAL IMAGING;127
15.7;VOLTAGE SENSITIVE RECORDINGS;128
15.8;INTRINSIC SIGNAL IMAGING;130
15.9;References;130
16;Presenting and Publishing Research Data;131
16.1;INTRODUCTION;131
16.2;OUTLETS FOR RESULTS;132
16.3;PUBLICATION IN PEER-REVIEWED JOURNALS;133
16.4;THE SPECTRUM OF JOURNALS;134
16.5;IMPACT FACTOR;134
16.6;WRITING THE PAPER;136
16.7;ASSESSMENT;137
16.8;FUTURE TRENDS;138
16.9;Ten key points;139
17;Analyzing Health Studies;140
17.1;OVERVIEW;140
17.2;TYPES OF DATA Numerical Data;140
17.3;Categorical Data;140
17.4;Data Structure;140
17.5;DESCRIBING DATA;141
17.6;Continuous Data;141
17.7;Categorical Data;142
17.8;ANALYZING DATA;142
17.9;Estimation;142
17.10;Hypothesis Testing;143
17.11;EXAMPLES;144
17.12;Two Independent Groups of Continuous Data;144
17.13;Two Paired Groups of Continuous Data;147
17.14;5.3 Two Independent Groups of Binary Data;149
17.15;Paired Binary Data;151
17.16;CONCLUSION;151
17.17;References;152
18;Future;153
19;Index;155