Ultrasound contrast agents (eBook)

Preface 5
Acknowledgments 7
Table of Contents 8
List of Contributors 10
Chapter 1 Towards Ultrasound Molecular Imaging 13
1.1 Introduction 13
1.2 Ultrasound molecular probes: new features of imaging 15
1.3 Proof of concept of molecular imaging with ultrasound 17
1.4 Targeted ultrasound contrast agents: new shuttles for drug delivery 18
1.5 Conclusions 20
References 21
Chapter 2 Use of Microbubbles as Ultrasound Contrast Agents for Molecular Imaging 24
2.1 Introduction 24
2.2 Molecular design 25
2.2.1 Contrast media 25
2.2.2 Targeting ligands 26
2.3 In vitro screening 28
2.3.1 Dynamic binding assay 28
2.3.2 Influence of ligand density on binding efficiency 28
2.4 In vivo proof of concept 30
2.4.1 Methodology 30
2.4.2 Tumor detection 31
2.5 Conclusions and prospects 33
References 33
Chapter 3 Design of Novel Polymer Shelled Ultrasound Contrast Agents: Towards an Ultrasound Triggered Drug Delivery 35
3.1 Introduction 35
3.2 Experimental section 37
3.2.1 Materials 37
3.2.2 Methods 37
Synthesis of PVA microballoons at pH5 and room temperature (MB5R) 37
Synthesis of PVA microballoons at pH 5 and 5.C (MB5C) 37
Synthesis of PVA microballoons at pH 2 and 5.C (MB2C) 37
Functionalization of MBs with NH2-PEG-NH2 38
Functionalization of MBs with RGD 38
Functionalization of MBs with L-cysteine(Cys-MB) 38
Synthesis of .- cyclodextrin (.CD) – functionalized MBs 38
Loading of NO on Cys-MB (NO-Cys-MB) 38
Synthesis of NPC functionalized PVA (NPC-PVA) 39
Synthesis of RGD functionalized PVA (RGD-PVA) 39
Synthesis of telechelic PVA 39
Synthesis of telechelic PVA hydrogels 39
Synthesis of RGD-PVA hydrogels 39
3.3 Results and discussion 40
3.3.1 Synthesis of PVA microballoons and surface characterization 40
3.3.2 Functionalization of MBs 41
3.3.3 RGD functionalized PVA, RGD-PVA 45
3.3.4 Synthesis of RGD-PVA hydrogels 46
3.4 Conclusions 48
References 48
Chapter 4 Lipidic Microbubble Targeting of Surface Proteins Using an in Vitro System 50
4.1 Introduction 50
4.2 Material and methods 52
4.2.1 Antibodies and reagents 52
4.2.2 Cell culture of U-937 for flow cytometry testing 52
4.2.3 TF receptor binding assay 52
4.2.4 Flow cytometry 53
4.2.5 Cell culture of U87-MG for perfusion testing 53
4.2.6 Maximum shear stress determination 54
4.2.7 Perfusion experiment with U87-MG 54
4.2.8 Image analysis 55
4.3 Results 55
4.3.1 Flow cytometry investigation of targeted microbubbles-cell interactions 55
4.3.2 Cell culture of U87-MG for tissue factor targeting in a perfused microchannel 56
4.3.3 Shear stress tolerance of U87-MG 57
4.3.4 Tissue factor targeting with microbubbles in a perfused microchannel 58
4.4 Conclusions 60
References 61
Chapter 5 Protein Interactions with Microballoons: Consequences for Biocompatibility and Application as Contrast Agents 62
5.1 Introduction 62
5.2 Experimental details 65
5.2.1 Human blood plasma 65
Buffers 65
Microballoons 65
Tissue culture medium 65
Proteins 65
1D Polyacrylamide gel electrophoesis (PAGE) 66
Mass spectrometry 66
Isothermal titration calorimetry (ITC) 66
5.3 Results and discussion 67
5.3.1 Dispersion of the microballoons in PBS buffer 67
5.3.2 Determination of the microballoon protein corona from plasma 67
5.3.3 Microballoon interaction with individual plasma proteins 70
5.3.4 Isothermal titration calorimetry 71
5.4 Discussion 72
5.5 Conclusions 73
References 74
Chapter 6 Polymer Based Biointerfaces: A Case Study on Devices for Theranostics and Tissue Engineering 76
6.1 Introduction 76
6.2 Materials and methods 77
6.2.1 Reagents 77
6.2.2 PVA hydrogels and MBs biocompatibility on NIH3T3 fibroblast mouse cell line 77
6.2.3 NIH3T3 fibroblasts and RAW 264.7 macrophage cultures 77
6.2.4 Microballoon suspension sterilization 78
6.2.5 NIH3T3 fibroblast and RAW 264.7 macrophage viability in the presence of PVA hydrogels, RGD modified PVA-hydrogels and PVA based MBs 78
6.2.6 NIH3T3 fibroblast and RAW264.7 macrophage proliferation on PVA hydrogels and after exposure to PVA based MBs 78
6.2.7 Phagocytosis of Microballoons 78
6.2.8 RAW 264.7 viability by live/dead assay 79
6.2.9 MB internalization studied by confocal laser scanning microscopy (CLSM) 79
6.3 Results and discussion 79
6.4 Conclusions 85
References 85
Chapter 7 Ultrasound Contrast Agent Microbubble Dynamics 87
7.1 Introduction 87
7.2 Dynamics of a free gas bubble 89
7.2.1 Linearized equations 90
7.3 Coated bubbles 91
7.3.1 Linearized equations 94
7.3.2 Optical and acoustical characterization 96
7.4 Bubble dynamics near a rigid wall 98
7.4.1 Method of images 98
7.4.2 Linearized equations 99
7.4.3 Observations 100
7.5 Conclusions 102
References 103
Chapter 8 Characterization of Acoustic Properties of PVA-Shelled Ultrasound Contrast Agents 106
8.1 Introduction 106
8.2 Materials and methods 107
8.2.1 Particle size distribution 107
8.2.2 Experimental setups 107
8.3 Results 109
8.3.1 Backscattered power spectra 109
8.3.2 Attenuation and dispersion 110
8.3.3 Ultrasound-induced fracture of PVA microbubbles 111
8.4 Conclusions 114
References 115
Chapter 9 Novel Characterization Techniques of Microballoons 116
9.1 Characterization of MBs by AFM 116
9.1.1 Experimental setup 117
9.1.2 Bubble bursting 118
9.1.3 Bubbles after bursting 119
9.1.4 Bubbles’ stiffness cycle dependence 119
9.1.5 Bubble stiffness 121
9.1.6 Bubble temperature dependence 121
9.1.8 Adhesion forces 123
9.1.9 Adhesion arrays 125
9.1.10 Conclusions 126
9.2 Characterization of MBs with STXM 126
References 132
Chapter 10 A Comparative Analysis of Contrast Enhanced Ultrasound Imaging Techniques 135
10.1 Introduction 135
10.2 Signal processing methods 137
10.2.1 Multi-pulse techniques 137
10.2.2 Chirp coding 138
10.2.3 Combining chirp coding with multi-pulse techniques 138
10.3 Simulation 139
10.3.1 Bubble response 139
10.3.2 Tissue propagation 140
10.3.3 A comprehensive simulation tool 140
Transmission 140
Propagation and interaction with microbubbles 141
Reception and post-processing 142
10.4 Evaluation metric 143
10.5 Results 144
10.5.1 Setup description 144
10.5.2 Analyses and comparison 145
10.6 Conclusions 147
References 147
Chapter 11 Remote Measurement of the Ultrasound Contrast Agent Concentration 149
11.1 Introduction 149
11.1.1 General approach 150
11.1.2 Tissue rejection 151
11.2 Concentration estimation 152
11.2.1 Feature vector 152
11.2.2 Non-linear regression 153
11.2.3 Support vector machine 154
11.2.4 Training set generation 156
11.3 The experimental set-up 156
11.4 Results 158
11.5 Conclusions 159
References 162
Chapter 12 Bubble Behavior Testing (BBT) System for Ultrasound Contrast Agent Characterization 163
12.1 Introduction 163
12.2 BBT system 165
12.2.1 Transducer front-end 165
12.2.2 Data management 168
12.2.3 Processing unit 168
12.2.4 Interfaces and synchronization capabilities 168
12.3 Observation of freely moving bubbles 169
12.3.1 Material and methods 169
12.3.2 Experimental results 170
12.3.3 Observation of bubble rupture 172
12.4 Observation of steady bubbles 174
12.4.1 Materials and methods 174
12.4.2 Experimental results 175
12.5 Conclusions 178
References 179
Chapter 13 A New Flexible Ultrasound Scanner Suitable for Research Topics 181
13.1 An industrial experience on UCM processing 181
13.2 The new scanner 182
13.2.1 General overview 183
Input Connector Switch board 185
Input TX-RX board 185
Input CW Clock board 188
Input Master Control board 189
Back-End Link Control board 189
Digital Image Processing board 189
Digital Elaboration Processing board 190
Digital Color-Flow Processing board 190
Back-End Scan Converter board 190
Digital Radio-Frequency Processing board 190
13.3 Conclusions 194
References 194
Index 195