Preface ix
Contributors xi
1 Overview of Genotyping 1
Monica Bayes and Ivo Glynne Gut
Introduction 1
Methods for interrogating SNPs 3
Commercial platforms for SNP genotyping 7
Practical recommendations 11
SNP databases 14
Methylation analysis 15
Copy number variation analysis 16
Second generation sequencing technologies 17
Conclusions 18
References 19
2 DNA Chip Analysis in Genome Discovery 24
Ross N. Nazar and Jane Robb
Introduction 24
Interrogating a genome 25
Cross-species hybridization 28
Comparative genomic hybridization and microarray-based genotyping 31
Barcodes, DNA microarrays and organism identification 34
Concluding remarks 36
References 37
3 qPCR, Theory, Reliability and Use in Molecular Analysis 43
Jamie Murphy and Stephen A. Bustin
Sample preparation 44
RNA quality 45
Reagents 46
Assay design 47
Transparency of published data 51
Further considerations 52
Conclusion 52
References 52
4 DNA Analysis in Droplet-Based Microfluidic Devices 56
Pinar Ozdemir and Yonghao Zhang
Introduction 56
Continuous-flow microPCR chips 58
PCR inhibition and carryover contamination 62
PCR in droplets 65
Conclusions 73
References 75
5 High-Resolution Melt Profiling 81
Steven F. Dobrowolski and Carl T. Wittwer
Introduction 81
Basic concepts of melt profiling 82
HRMP and polymerase chain reaction 84
DNA specimens and HRMP 89
Determining a temperature window for melting 90
Dyes and platforms for melt profiling 92
Scanning PCR products for sequence variation 93
Genotyping with high-resolution melt profiling 102
Other applications of HRMP 108
Final notes 109
References 110
6 Massively Parallel Sequencing 114
Tracy Tucker, Marco Marra and Jan M. Friedman
Sanger sequencing 114
Massively parallel sequencing 114
Commercially available massively parallel sequencers 115
Future technologies 122
Paired-end or mate-paired reads 123
Target-enrichment strategies for MPS 124
Applications of MPS 125
Summary 131
References 132
7 Aptamers for Analysis: Nucleic Acids Ligands in the Post-Genomic Era 135
Pedro Nadal, Alessandro Pinto, Marketa Svobodova and Ciara K. O'Sullivan
Introduction 135
SELEX 136
Aptamers in analysis 147
Imaging with aptamers 157
Conclusions, outlooks and perspectives 160
References 161
8 Use of Nanotechnology for Enhancing of Cancer Biomarker Discovery and Analysis: A Molecular Approach 175
Farid E. Ahmed
Introduction 175
Proteomics and nanotechnology 176
Nanoscale multicomponent separation 176
Nanoscale protein detection strategies 178
Surface-enhanced Raman scattering (SERS) 182
References 188
9 Chip-Based Proteomics 193
Julian Bailes, Andrew Milnthorpe, Sandra Smieszek and Mikhail Soloviev
Introduction 193
Lab-on-a-chip 194
Arrays 196
Chip-based mass spectrometry 203
Surface plasmon resonance (SPR) and quartz crystal microbalance (QCM) chip instruments 205
Microfluidics 207
Conclusion 209
References 211
10 Antibody Microarrays in Proteome Profiling 219
Mohamed Saiel Saeed Alhamdani and Jorg D. Hoheisel
Introduction 219
Technical aspects 220
Antibody array applications 226
Summary 229
Acknowledgements 229
References 229
11 Biomarker Detection and Molecular Profiling by Multiplex Microbead Suspension Array Based Immunoproteomics 244
V. V. Krishhan, Imran H. Khan and Paul A. Luciw
Introduction 244
Principles of microbead-based multiplexing 245
Experimental aspects of the multiplex microbead assay 247
Multiplex microbead assay design and comparison with other methods 250
Applications of the multiplex microbead assay system for biomedical research and clinical studies 252
Selected investigational fields for multiplex analysis and examples of applications 255
Challenges and current limitations 255
Summary and future directions 259
Acknowledgements 261
References 261
12 Mass Spectrometry in Metabolomics 271
William J. Griffiths and Yuqin Wang
Introduction 271
Sample collection and preparation 272
Data acquisition 274
Data analysis 279
Applications 283
Conclusion 294
Acknowledgement 294
References 295
Index 299
