Detection of Highly Dangerous Pathogens: Microarray Methods for BSL 3 and BSL 4 Agents

Detection of Highly Dangerous Pathogens: Microarray Methods for the Detection of BSL 3 and BSL 4 Agents......Page 1
Contents......Page 7
This Publication is Supported by COST......Page 13
Preface......Page 15
List of Contributors......Page 17
1.1 Introduction to Microarray Technology......Page 21
1.2.1 Probes......Page 22
1.2.1.3 Oligonucleotide Probes......Page 23
1.2.2 Substrates for Printing......Page 26
1.2.2.6 Probe Spacers......Page 29
1.2.3.1 Target Amplifications and Sensitivity Issues......Page 30
1.2.3.3 Hybridization and Wash Conditions......Page 31
1.2.4.2 In Situ Synthesis......Page 32
1.2.5.2 Planar-Waveguide Technology (PWT)......Page 34
1.2.5.4 Three-Dimensional Microarray Formats......Page 35
1.2.6 Marker Genes Used on MDMs......Page 36
1.3 Analysis and Quality Control Aspects......Page 37
1.4.2 Comparative Genomic Hybridization (CGH)......Page 38
1.4.3 Generic or Universal Microarrays......Page 39
1.4.4 Microarrays for Sequence Analysis......Page 40
1.4.5 Microbial Diagnostic Microarrays (MDMs)......Page 41
References......Page 42
Part I: Methods......Page 55
2.1 Introduction......Page 57
2.2.2 Φ29 Amplification......Page 58
2.2.3 Klenow Amplification/Labeling......Page 59
2.2.4 Probe and Slide Preparation......Page 60
2.2.5 Slide Processing Protocol (for Amino Surfaces)......Page 61
2.2.6 Hybridization and Slide Washing......Page 62
2.4 Conclusions......Page 64
References......Page 66
3.1 Introduction......Page 67
3.2 Probe Design......Page 70
3.3 Slide Preparation (Spotting)......Page 71
3.5 DNA Extraction and PCR Amplification of the Targeted Gene......Page 72
3.7 Labeling......Page 73
3.8 Hybridization and Slide Washing......Page 74
3.11 Microarray for Detection of Pathogenic Bacteria......Page 75
References......Page 76
4.1 Introduction......Page 79
4.2.2 Definition of the Optimal Probe Length......Page 80
4.2.3 Virtual Assessment of Array Performances (in Silico Experiments)......Page 81
4.2.4 Array Manufacturing and Hybridization (Wet-Lab Experiments)......Page 83
References......Page 84
5.1 Introduction......Page 87
5.2 Soft Lithography......Page 88
5.3 Photolithography......Page 90
5.4.1 Micro-Spotting......Page 91
5.4.2 Ink-Jet Printing......Page 92
5.5 Lithography with AFM......Page 94
5.5.1 Dip-Pen Lithography with AFM......Page 95
5.5.2 cAFM Lithography......Page 96
5.5.3 Nanoshaving and Nanografting......Page 97
5.6 Conclusions......Page 100
References......Page 101
6.1 Introduction......Page 105
6.2.1 Glass......Page 106
6.2.3 Gold......Page 110
6.2.5 QDs......Page 111
6.3 Immobilization......Page 114
References......Page 119
Part II: Identification......Page 125
7.2 Chip Design/Array Description......Page 127
7.3.1 Amplification......Page 128
7.3.2 Short Protocol for Hybridization and Labeling......Page 129
7.5 Conclusions......Page 131
8.2 Principle of Applying Microarray Technology for Virus Detection and Identification......Page 133
8.4 Advantages and Drawbacks of Using the Microarray Technique......Page 135
8.5 Key Factors for Development of a Microarray-Based Test......Page 136
8.7 Flavivirus Microarray......Page 137
8.8 Hemorrhagic Fever Viruses......Page 141
References......Page 143
9.2 Francisella Genomotyping......Page 145
9.3 Brucella Genomotyping......Page 148
9.4 Conclusions......Page 150
References......Page 151
Part III: Typing......Page 153
10.1.1 Real-Time Polymerase Chain Reaction: More Than a Simple Polymerase Chain Reaction......Page 155
10.2.1.1 Use of TaqMan LNA Probes for the Typing of Brucella suis Subspecies......Page 156
10.2.3 Typing Large Number of SNPs by Ligation Detection Reaction......Page 159
10.2.3.1 Use of LDR Probes and Low-Density Microarrays for the Multiplex Diagnostic of BSL3 Bacteria......Page 162
10.3 Conclusions......Page 164
References......Page 165
11.1 Introduction......Page 167
11.3 Determination of the MICs of Different Antibiotics for the B. anthracis Transconjugants......Page 168
11.4 Detection of Antibiotic Resistance Genes in B. anthracis by Microchip-Based Hybridization System (ArrayTube)......Page 169
11.5 Conclusions......Page 170
References......Page 171
Part IV: Quality Control......Page 173
12.1 Introduction......Page 175
12.2.2 Design and Printing of Oligonucleotide Probes......Page 177
12.2.4 Hybridization of Labeled Targets and Data Processing......Page 179
12.3.1 Random Amplification and Hybridization of Cy3-labeled Pathogen DNA Targets......Page 180
12.3.2 Quality Assessment of Intra- and Inter-Operator Variation......Page 181
12.3.2.1 Sources of Intra-Operator Experimental Variation......Page 182
12.3.2.2 Sources of Inter-Operator Experimental Variation......Page 184
References......Page 185
Index......Page 189