Dual Ultrasound and Photoacoustic Tracking of Magnetically Driven Micromotors: From In Vitro to In Vivo

The fast evolution of medical micro- and nanorobots in the endeavor to perform non-invasive medical operations in living organisms has boosted the use of diverse medical imaging techniques in the last years. Among those techniques, photoacoustic imaging (PAI), considered a functional technique, has shown to be promising for the visualization of micromotors in deep tissue with high spatiotemporal resolution as it possesses the molecular specificity of optical methods and the penetration depth of ultrasound. However, the precise maneuvering and function's control of medical micromotors, in particular in living organisms, require both anatomical and functional imaging feedback. Therefore, herein, the use of high-frequency ultrasound and PAI is reported to obtain anatomical and molecular information, respectively, of magnetically-driven micromotors in vitro and under ex vivo tissues. Furthermore, the steerability of the micromotors is demonstrated by the action of an external magnetic field into the uterus and bladder of living mice in real-time, being able to discriminate the micromotors’ signal from one of the endogenous chromophores by multispectral analysis. Finally, the successful loading and release of a model cargo by the micromotors toward non-invasive in vivo medical interventions is demonstrated. © 2021 The Authors. Advanced Healthcare Materials published by Wiley-VCH Gmb

Medical Imaging of Microrobots: Toward In Vivo Applications

Medical microrobots (MRs) have been demonstrated for a variety of non-invasive biomedical applications, such as tissue engineering, drug delivery, and assisted fertilization, among others. However, most of these demonstrations have been carried out in in vitro settings and under optical microscopy, being significantly different from the clinical practice. Thus, medical imaging techniques are required for localizing and tracking such tiny therapeutic machines when used in medical-relevant applications. This review aims at analyzing the state of the art of microrobots imaging by critically discussing the potentialities and limitations of the techniques employed in this field. Moreover, the physics and the working principle behind each analyzed imaging strategy, the spatiotemporal resolution, and the penetration depth are thoroughly discussed. The paper deals with the suitability of each imaging technique for tracking single or swarms of MRs and discusses the scenarios where contrast or imaging agent's inclusion is required, either to absorb, emit, or reflect a determined physical signal detected by an external system. Finally, the review highlights the existing challenges and perspective solutions which could be promising for future in vivo applications

Electronically integrated microcatheters based on self-assembling polymer films

Existing electronically integrated catheters rely on the manual assembly of separate components to integrate sensing and actuation capabilities. This strongly impedes their miniaturization and further integration. Here, we report an electronically integrated self-assembled microcatheter. Electronic components for sensing and actuation are embedded into the catheter wall through the self-assembly of photolithographically processed polymer thin films. With a diameter of only about 0.1 mm, the catheter integrates actuated digits for manipulation and a magnetic sensor for navigation and is capable of targeted delivery of liquids. Fundamental functionalities are demonstrated and evaluated with artificial model environments and ex vivo tissue. Using the integrated magnetic sensor, we develop a strategy for the magnetic tracking of medical tools that facilitates basic navigation with a high resolution below 0.1 mm. These highly flexible and microsized integrated catheters might expand the boundary of minimally invasive surgery and lead to new biomedical applications. Copyright © 2021 The Authors, some rights reserved

Medical Imaging of Magnetic Micromotors Through Scattering Tissues

Micro- and nanorobots (MNRs) are small autonomous devices capable of performing complex tasks and have been demonstrated for a variety of non-invasive biomedical applications, such as tissue engineering, drug delivery or assisted fertilization. However, translating such approaches to an in vivo environment is critical. Current imaging techniques do not allow localization and tracking of single or few micromotors at high spatiotemporal resolution in deep tissue. This thesis addresses some of these limitations, by exploring the use of two optical-based techniques (IR and photoacoustic imaging (PAI)) and a combination of both US and PAI. First, we employ an IR imaging setup to visualize mobile reflective micromotors under scattering phantoms and ex vivo mouse skull tissues, without using any labels. The reflective micromotor reflects more than tenfold the light intensity of a simple particle. However, the achieved penetration depth was ca. 100 μm (when using ex vivo tissues), limiting this technique to superficial biomedical applications. In this regard, PAI plays a role that combines the advantages of US such as penetration depth and real-time imaging with the molecular specificity of optics. For the first time, in this thesis, this method is evaluated for dynamic process monitoring, in particular for tracking single micromotor in real-time below ~1 cm deep phantom and ex vivo tissue. However, the precise function control of MNRs in living organisms, demand the combination of both anatomical and functional imaging methods. Therefore, in the end, we report the use of a hybrid US and PA system for the real-time tracking of magnetically driven micromotors (single and swarms) in phantoms, ex vivo, and in vivo (in mice bladder and uterus), envisioning their application for targeted drug-delivery. This achievement is of great importance and opens the possibilities to employ medical micromotors in a living organism and perform a medical task while being externally controlled and monitored.:ABSTRACT 1 1 INTRODUCTION 5 1.1 Motivation 5 1.2 Background 7 1.2.1 Microrobotics 7 1.2.2 Medical Imaging 9 1.3 Objectives and Structure of Thesis 12 2 FUNDAMENTALS 15 2.1 Optical Imaging 15 2.1.1. Reflection-based Imaging 17 2.1.2. Fluorescence-based Imaging 18 2.1.1 Light-Tissue Interaction 20 2.2 Photoacoustic Imaging 23 2.2.1 Theory 23 2.2.2 Implementation 25 2.3 Ultrasound Imaging 26 2.3.1 Theory 26 2.3.2 Implementation 28 3 MATERIALS AND METHODS 30 3.1 Fabrication of Magnetic Micropropellers 30 3.1.1 3D Laser Lithography of Polymeric Resin 30 3.1.2 Self-assembly of SiO2 Particles 31 3.1.3 Electron Beam Evaporation 32 3.1.4 Surface Functionalization 33 3.2 Fabrication of Phantom Tissue and Microfluidic Channels 34 3.2.1 Fabrication of PDMS-Glycerol Phantom 34 3.2.2 Fabrication of Agarose Phantom 35 3.2.3 Phantom based on Ex vivo Tissues (Chicken Breast and Mice Skull) 36 3.2.4 Microfluidic Channel Platform 37 3.3 Sample Characterization 38 3.3.1 Optical Microscopy 38 3.3.2 Scanning Electron Microscopy 38 3.4 Magnetic Actuation 39 3.4.1 Magnetic Force 39 3.4.2 Magnetic Torque 39 3.5 Ethic Statement for Mice Experiments 41 4 OPTICAL IMAGING OF MICROROBOTS 42 4.1 Concept of Reflective Micromotors 42 4.2 Fabrication of Reflective Micromotors 44 4.3 IR Imaging Actuation Setup 45 4.4 Actuation and Propulsion Performance below Phantom 47 4.5 Actuation and Propulsion Performance below Ex Vivo Skull Tissue 50 4.6 Actuation and Propulsion Performance in Blood 51 5 PHOTOACOUSTIC IMAGING OF MICROROBOTS 55 5.1 Absorbers for Deep Tissue Imaging 55 5.2 Absorber Micromotor Design and Fabrication 56 5.3 Photoacoustic Imaging Setup 58 5.4 Actuation Performance below Phantom Tissue 60 5.5 Actuation Performance below Ex Vivo Tissue 65 6 HYBRID ULTRASOUND AND PHOTOACOUSTIC IMAGING 67 6.1 Hybrid Ultrasound/Photoacoustic System 68 6.2 Fabrication and Characterization of Micromotors 69 6.3 Actuation and Propulsion Performance below Phantom 69 6.4 Actuation and Propulsion Performance below Ex Vivo Tissues 71 6.5 Actuation and Propulsion Performance in Mice 72 6.5.1 Swimming of Micromotors in Bladder 72 6.5.2 Actuation of Micromotors in Uterus 74 6.5.3 3D Multispectral Imaging 76 6.5.4 Towards Targeted Drug Delivery 77 7 SUMMARY AND PERSPECTIVES 80 7.1 Summary 80 7.2 Future Perspectives 83 7.2.1 Contrats Enhancing Labels 84 7.2.2 Novel Imaging Concepts 85 8 REFERENCES 88 9 APPENDIX 105 List of Figures 105 List of Tables 107 Abbreviations 108 List of Publications 109 Acknowledgements 110 Selbstständigkeitserklärung 111 Curriculum Vitae 11

Medical Imaging of Magnetic Micromotors Through Scattering Tissues

Micro- and nanorobots (MNRs) are small autonomous devices capable of performing complex tasks and have been demonstrated for a variety of non-invasive biomedical applications, such as tissue engineering, drug delivery or assisted fertilization. However, translating such approaches to an in vivo environment is critical. Current imaging techniques do not allow localization and tracking of single or few micromotors at high spatiotemporal resolution in deep tissue. This thesis addresses some of these limitations, by exploring the use of two optical-based techniques (IR and photoacoustic imaging (PAI)) and a combination of both US and PAI. First, we employ an IR imaging setup to visualize mobile reflective micromotors under scattering phantoms and ex vivo mouse skull tissues, without using any labels. The reflective micromotor reflects more than tenfold the light intensity of a simple particle. However, the achieved penetration depth was ca. 100 μm (when using ex vivo tissues), limiting this technique to superficial biomedical applications. In this regard, PAI plays a role that combines the advantages of US such as penetration depth and real-time imaging with the molecular specificity of optics. For the first time, in this thesis, this method is evaluated for dynamic process monitoring, in particular for tracking single micromotor in real-time below ~1 cm deep phantom and ex vivo tissue. However, the precise function control of MNRs in living organisms, demand the combination of both anatomical and functional imaging methods. Therefore, in the end, we report the use of a hybrid US and PA system for the real-time tracking of magnetically driven micromotors (single and swarms) in phantoms, ex vivo, and in vivo (in mice bladder and uterus), envisioning their application for targeted drug-delivery. This achievement is of great importance and opens the possibilities to employ medical micromotors in a living organism and perform a medical task while being externally controlled and monitored.:ABSTRACT 1 1 INTRODUCTION 5 1.1 Motivation 5 1.2 Background 7 1.2.1 Microrobotics 7 1.2.2 Medical Imaging 9 1.3 Objectives and Structure of Thesis 12 2 FUNDAMENTALS 15 2.1 Optical Imaging 15 2.1.1. Reflection-based Imaging 17 2.1.2. Fluorescence-based Imaging 18 2.1.1 Light-Tissue Interaction 20 2.2 Photoacoustic Imaging 23 2.2.1 Theory 23 2.2.2 Implementation 25 2.3 Ultrasound Imaging 26 2.3.1 Theory 26 2.3.2 Implementation 28 3 MATERIALS AND METHODS 30 3.1 Fabrication of Magnetic Micropropellers 30 3.1.1 3D Laser Lithography of Polymeric Resin 30 3.1.2 Self-assembly of SiO2 Particles 31 3.1.3 Electron Beam Evaporation 32 3.1.4 Surface Functionalization 33 3.2 Fabrication of Phantom Tissue and Microfluidic Channels 34 3.2.1 Fabrication of PDMS-Glycerol Phantom 34 3.2.2 Fabrication of Agarose Phantom 35 3.2.3 Phantom based on Ex vivo Tissues (Chicken Breast and Mice Skull) 36 3.2.4 Microfluidic Channel Platform 37 3.3 Sample Characterization 38 3.3.1 Optical Microscopy 38 3.3.2 Scanning Electron Microscopy 38 3.4 Magnetic Actuation 39 3.4.1 Magnetic Force 39 3.4.2 Magnetic Torque 39 3.5 Ethic Statement for Mice Experiments 41 4 OPTICAL IMAGING OF MICROROBOTS 42 4.1 Concept of Reflective Micromotors 42 4.2 Fabrication of Reflective Micromotors 44 4.3 IR Imaging Actuation Setup 45 4.4 Actuation and Propulsion Performance below Phantom 47 4.5 Actuation and Propulsion Performance below Ex Vivo Skull Tissue 50 4.6 Actuation and Propulsion Performance in Blood 51 5 PHOTOACOUSTIC IMAGING OF MICROROBOTS 55 5.1 Absorbers for Deep Tissue Imaging 55 5.2 Absorber Micromotor Design and Fabrication 56 5.3 Photoacoustic Imaging Setup 58 5.4 Actuation Performance below Phantom Tissue 60 5.5 Actuation Performance below Ex Vivo Tissue 65 6 HYBRID ULTRASOUND AND PHOTOACOUSTIC IMAGING 67 6.1 Hybrid Ultrasound/Photoacoustic System 68 6.2 Fabrication and Characterization of Micromotors 69 6.3 Actuation and Propulsion Performance below Phantom 69 6.4 Actuation and Propulsion Performance below Ex Vivo Tissues 71 6.5 Actuation and Propulsion Performance in Mice 72 6.5.1 Swimming of Micromotors in Bladder 72 6.5.2 Actuation of Micromotors in Uterus 74 6.5.3 3D Multispectral Imaging 76 6.5.4 Towards Targeted Drug Delivery 77 7 SUMMARY AND PERSPECTIVES 80 7.1 Summary 80 7.2 Future Perspectives 83 7.2.1 Contrats Enhancing Labels 84 7.2.2 Novel Imaging Concepts 85 8 REFERENCES 88 9 APPENDIX 105 List of Figures 105 List of Tables 107 Abbreviations 108 List of Publications 109 Acknowledgements 110 Selbstständigkeitserklärung 111 Curriculum Vitae 11

Dual Ultrasound and Photoacoustic Tracking of Magnetically Driven Micromotors: From In Vitro to In Vivo

The fast evolution of medical micro- and nanorobots in the endeavor to perform non-invasive medical operations in living organisms has boosted the use of diverse medical imaging techniques in the last years. Among those techniques, photoacoustic imaging (PAI), considered a functional technique, has shown to be promising for the visualization of micromotors in deep tissue with high spatiotemporal resolution as it possesses the molecular specificity of optical methods and the penetration depth of ultrasound. However, the precise maneuvering and function's control of medical micromotors, in particular in living organisms, require both anatomical and functional imaging feedback. Therefore, herein, the use of high-frequency ultrasound and PAI is reported to obtain anatomical and molecular information, respectively, of magnetically-driven micromotors in vitro and under ex vivo tissues. Furthermore, the steerability of the micromotors is demonstrated by the action of an external magnetic field into the uterus and bladder of living mice in real-time, being able to discriminate the micromotors’ signal from one of the endogenous chromophores by multispectral analysis. Finally, the successful loading and release of a model cargo by the micromotors toward non-invasive in vivo medical interventions is demonstrated

Real-Time IR Tracking of Single Reflective Micromotors through Scattering Tissues

Medical micromotors have the potential to lead to a paradigm shift in future biomedicine, as they may perform active drug delivery, microsurgery, tissue engineering, or assisted fertilization in a minimally invasive manner. However, the translation to clinical treatment is challenging, as many applications of single or few micromotors require real-time tracking and control at high spatiotemporal resolution in deep tissue. Although optical techniques are a popular choice for this task, absorption and strong light scattering lead to a pronounced decrease of the signal-to-noise ratio with increasing penetration depth. Here, a highly reflective micromotor is introduced which reflects more than tenfold the light intensity of simple gold particles and can be precisely navigated by external magnetic fields. A customized optical IR imaging setup and an image correlation technique are implemented to track single micromotors in real-time and label-free underneath phantom and ex vivo mouse skull tissues. As a potential application, the micromotors speed is recorded when moving through different viscous fluids to determine the viscosity of diverse physiological fluids toward remote cardiovascular disease diagnosis. Moreover, the micromotors are loaded with a model drug to demonstrate their cargotransport capability. The proposed reflective micromotor is suitable as theranostic tool for sub-skin or organ-on-a-chip applications

Real-Time IR Tracking of Single Reflective Micromotors through Scattering Tissues

Medical micromotors have the potential to lead to a paradigm shift in future biomedicine, as they may perform active drug delivery, microsurgery, tissue engineering, or assisted fertilization in a minimally invasive manner. However, the translation to clinical treatment is challenging, as many applications of single or few micromotors require real-time tracking and control at high spatiotemporal resolution in deep tissue. Although optical techniques are a popular choice for this task, absorption and strong light scattering lead to a pronounced decrease of the signal-to-noise ratio with increasing penetration depth. Here, a highly reflective micromotor is introduced which reflects more than tenfold the light intensity of simple gold particles and can be precisely navigated by external magnetic fields. A customized optical IR imaging setup and an image correlation technique are implemented to track single micromotors in real-time and label-free underneath phantom and ex vivo mouse skull tissues. As a potential application, the micromotors speed is recorded when moving through different viscous fluids to determine the viscosity of diverse physiological fluids toward remote cardiovascular disease diagnosis. Moreover, the micromotors are loaded with a model drug to demonstrate their cargo-transport capability. The proposed reflective micromotor is suitable as theranostic tool for sub-skin or organ-on-a-chip applications. © 2019 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Real‐Time IR Tracking of Single Reflective Micromotors through Scattering Tissues

Medical micromotors have the potential to lead to a paradigm shift in future biomedicine, as they may perform active drug delivery, microsurgery, tissue engineering, or assisted fertilization in a minimally invasive manner. However, the translation to clinical treatment is challenging, as many applications of single or few micromotors require real-time tracking and control at high spatiotemporal resolution in deep tissue. Although optical techniques are a popular choice for this task, absorption and strong light scattering lead to a pronounced decrease of the signal-to-noise ratio with increasing penetration depth. Here, a highly reflective micromotor is introduced which reflects more than tenfold the light intensity of simple gold particles and can be precisely navigated by external magnetic fields. A customized optical IR imaging setup and an image correlation technique are implemented to track single micromotors in real-time and label-free underneath phantom and ex vivo mouse skull tissues. As a potential application, the micromotors speed is recorded when moving through different viscous fluids to determine the viscosity of diverse physiological fluids toward remote cardiovascular disease diagnosis. Moreover, the micromotors are loaded with a model drug to demonstrate their cargo-transport capability. The proposed reflective micromotor is suitable as theranostic tool for sub-skin or organ-on-a-chip applications. © 2019 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim