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Optical microscopy, providing valuable biomedical insights at the cellular and organelle levels, has been widely recognized as an enabling technology. Mainstream optical microscopy technologies, including single-/multi-photon fluorescence microscopy and OCT, have demonstrated extraordinary sensitivities to fluorescence and optical scattering contrasts, respectively. However, the optical absorption contrast of biological tissues, which encodes essential physiological/pathological information, has not yet been fully assessable. The emergence of biomedical photoacoustics has led to a new branch of optical microscopy--OR-PAM. As a valuable complement to existing optical microscopy technologies, OR-PAM detects optical absorption contrasts with exquisite sensitivity (i.e., 100%). Combining OR-PAM with fluorescence microscopy or optical-scattering-based OCT (or both) provides comprehensive optical properties of biological tissues. Moreover, OR-PAM encodes optical absorption into acoustic waves, in contrast to the pure optical processes in fluorescence microscopy and OCT, and thus provides background-free detection. The acoustic detection in OR-PAM mitigates the impacts of optical scattering on signal degradation and naturally eliminates possible interferences (i.e., crosstalks) between excitation and detection, which is a common problem in fluorescence microscopy due to the overlap between the excitation and fluorescence spectra and imperfect extinction of the filter. Unique for high-resolution imaging of optical absorption, OR-PAM has demonstrated broad biomedical applications in fields such as neurology, ophthalmology, vascular biology, and dermatology. My doctoral research focuses on developments and biomedical applications of OR-PAM. The first part of my dissertation discusses the development of three generations of OR-PAM towards high-resolution, high-sensitivity, high-speed, and wide FOV in vivo imaging. In this section, I provide a comprehensive description of OR-PAM, including the principle, system design, system configuration, experimental procedures, laser safety, functional imaging scheme, and example biomedical applications at a variety of in vivo anatomical sites (i.e., skins, eyes and brains). The second part of my dissertation focuses on the application of OR-PAM in vascular biology, with an emphasis on neovascularization. In this section, I demonstrate longitudinal OR-PAM monitoring of the morphological (i.e., vessel diameter, length, tortuosity and volume) and functional (i.e., sO2) changes of angiogenic microenvironment at the capillary level, in both a non-disease TetON-HIF-1 transgenic mouse model and a cancer xenograft model in mouse ear. The last part of my dissertation focuses on the application of OR-PAM in neurology, with an emphasis on cortical stimulation, Alzheimer's disease, and ischemic stroke. In this section, I use label-free OR-PAM for both acute monitoring of microvascular responses to direct electrical stimulations of the mouse somatosensory cortex through a cranial opening and longitudinal monitoring of the morphological and functional changes of cortical vasculature in a transient middle cerebral artery occlusion mouse model. I also explore the potential of OR-PAM for transcranial monitoring of amyloid plaque growth in an AD mouse model.
This open access book provides a comprehensive overview of the application of the newest laser and microscope/ophthalmoscope technology in the field of high resolution imaging in microscopy and ophthalmology. Starting by describing High-Resolution 3D Light Microscopy with STED and RESOLFT, the book goes on to cover retinal and anterior segment imaging and image-guided treatment and also discusses the development of adaptive optics in vision science and ophthalmology. Using an interdisciplinary approach, the reader will learn about the latest developments and most up to date technology in the field and how these translate to a medical setting. High Resolution Imaging in Microscopy and Ophthalmology – New Frontiers in Biomedical Optics has been written by leading experts in the field and offers insights on engineering, biology, and medicine, thus being a valuable addition for scientists, engineers, and clinicians with technical and medical interest who would like to understand the equipment, the applications and the medical/biological background. Lastly, this book is dedicated to the memory of Dr. Gerhard Zinser, co-founder of Heidelberg Engineering GmbH, a scientist, a husband, a brother, a colleague, and a friend.
This open access book provides a comprehensive overview of the application of the newest laser and microscope/ophthalmoscope technology in the field of high resolution imaging in microscopy and ophthalmology. Starting by describing High-Resolution 3D Light Microscopy with STED and RESOLFT, the book goes on to cover retinal and anterior segment imaging and image-guided treatment and also discusses the development of adaptive optics in vision science and ophthalmology. Using an interdisciplinary approach, the reader will learn about the latest developments and most up to date technology in the field and how these translate to a medical setting. High Resolution Imaging in Microscopy and Ophthalmology - New Frontiers in Biomedical Optics has been written by leading experts in the field and offers insights on engineering, biology, and medicine, thus being a valuable addition for scientists, engineers, and clinicians with technical and medical interest who would like to understand the equipment, the applications and the medical/biological background. Lastly, this book is dedicated to the memory of Dr. Gerhard Zinser, co-founder of Heidelberg Engineering GmbH, a scientist, a husband, a brother, a colleague, and a friend.
Brings a fresh point of view to the current state of correlative imaging and the future of the field This book provides contributions from international experts on correlative imaging, describing their vision of future developments in the field based on where it is today. Starting with a brief historical overview of how the field evolved, it presents the latest developments in microscopy that facilitate the correlative workflow. It also discusses the need for an ideal correlative probe, applications in proteomic and elemental analysis, interpretation methods, and how correlative imaging can incorporate force microscopy, soft x-ray tomography, and volume electron microscopy techniques. Work on placing individual molecules within cells is also featured. Correlative Imaging: Focusing on the Future offers in-depth chapters on: correlative imaging from an LM perspective; the importance of sample processing for correlative imaging; correlative light and volume EM; correlation with scanning probe microscopies; and integrated microscopy. It looks at: cryo-correlative microscopy; correlative cryo soft X-ray imaging; and array tomography. Hydrated-state correlative imaging in vacuo, correlating data from different imaging modalities, and big data in correlative imaging are also considered. Brings a fresh view to one of the hottest topics within the imaging community: the correlative imaging field Discusses current research and offers expert thoughts on the field’s future developments Presented by internationally-recognized editors and contributors with extensive experience in research and applications Of interest to scientists working in the fields of imaging, structural biology, cell biology, developmental biology, neurobiology, cancer biology, infection and immunity, biomaterials and biomedicine Part of the Wiley–Royal Microscopical Society series Correlative Imaging: Focusing on the Future will appeal to those working in the expanding field of the biosciences, correlative microscopy and related microscopic areas. It will also benefit graduate students working in microscopy, as well as anyone working in the microscopy imaging field in biomedical research.
This book offers an overview of imaging techniques used to investigate cells and tissue in their native environment. It covers the range of imaging approaches used, as well as the application of those techniques to the study of biological processes in cells and whole tissues within living organisms.
Photoacoustics promises to revolutionize medical imaging and may well make as dramatic a contribution to modern medicine as the discovery of the x-ray itself once did. Combining electromagnetic and ultrasonic waves synergistically, photoacoustics can provide deep speckle-free imaging with high electromagnetic contrast at high ultrasonic resolution and without any health risk. While photoacoustic imaging is probably the fastest growing biomedical imaging technology, this book is the first comprehensive volume in this emerging field covering both the physics and the remarkable noninvasive applications that are changing diagnostic medicine. Bringing together the leading pioneers in this field to write about their own work, Photoacoustic Imaging and Spectroscopy is the first to provide a full account of the latest research and developing applications in the area of biomedical photoacoustics. Photoacoustics can provide functional sensing of physiological parameters such as the oxygen saturation of hemoglobin. It can also provide high-contrast functional imaging of angiogenesis and hypermetabolism in tumors in vivo. Discussing these remarkable noninvasive applications and so much more, this reference is essential reading for all researchers in medical imaging and those clinicians working at the cutting-edge of modern biotechnology to develop diagnostic techniques that can save many lives and just as importantly do no harm.
This is the second volume of Advances in Acoustic Microscopy. It continues the aim of presenting applications and developments of techniques that are related to high-resolution acoustic imaging. We are very grateful to the authors who have devoted considerable time to preparing these chapters, each of which describes a field of growing importance. Laboratories that have high-performance acoustic microscopes are frequently asked to examine samples for which the highest available resolution is not necessary, and the ability to penetrate opaque layers is more significant. Such applications can be thought of as bridging the gap be tween acoustic microscopy at low gigahertz frequencies, and on the one hand nondestructive testing of materials at low megahertz frequencies and on the other hand medical ultrasonic imaging at low megahertz frequencies. Commercial acoustic microscopes are becoming increasingly available and popular for such applications. We are therefore delighted to be able to begin the volume with chapters from each of those two fields. The first chapter, by Gabriele Pfannschmidt, describes uses of acoustic microscopy in the semiconductor industry. It provides a splendid balance to the opening chapter of Volume 1, which came from a national research center, being written from within a major European electronics industry itself. Dr Pfann schmidt describes the use of two quite different types of acoustic microscopes, and points out the advantages of each for specific purposes.
Optical microscopy, thanks to the noninvasive nature of its measurement, takes a crucial role across science and engineering, and is particularly important in biological and medical fields. To meet ever increasing needs on its capability for advanced scientific research, even more diverse microscopic imaging techniques and their upgraded versions have been intensively developed over the past two decades. However, advanced microscopy development faces major challenges including super-resolution (beating the diffraction limit), imaging penetration depth, imaging speed, and label-free imaging. This dissertation aims to study high numerical aperture (NA) imaging methods proposed to tackle these imaging challenges. The dissertation first details advanced optical imaging theory needed to analyze the proposed high NA imaging methods. Starting from the classical scalar theory of optical diffraction and (partially coherent) image formation, the rigorous vectorial theory that handles the vector nature of light, i.e., polarization, is introduced. New sign conventions for polarization ray tracing based on a generalized Jones matrix formalism are established to facilitate the vectorial light propagation with physically consistent outcomes. The first high NA microscopic imaging of interest is wide-field oblique plane microscopy (OPM) for high-speed deep imaging. It is a simple, real-time imaging technique recently developed to access any inclined cross-section of a thick sample. Despite its experimental demonstration implemented by tilted remote focusing, the optical resolution of the method has not been fully understood. The anisotropic resolving power in high NA OPM is rigorously investigated and interpreted by deriving the vectorial point spread function (PSF) and vectorial optical transfer function (OTF). Next, OPM is combined with stochastic optical reconstruction microscopy (STORM) to achieve super-resolution deep imaging. The proposed method, termed obliqueSTORM, together with oblique lightsheet illumination paves the way for deeper penetration readily available in localization-based super-resolution microscopy. The key performance metrics of obliqueSTORM, quantitative super-resolution andaxial depth of field, are studied. obliqueSTORM could achieve sub-50-nm resolution with a penetration depth of tens of microns for biological samples. The last part of the thesis covers the development of nonparaxial imaging theory of high NA differential phase contrast (DPC) microscopy for high resolution quantitative phase imaging. The phase retrieval in conventional optical DPC microscopy relies on the paraxial transmission cross-coefficient (TCC) model. However, this paraxial model becomes invalid in high NA DPC imaging. Formulated here is a more advanced nonparaxial TCC model that considers the nonparaxial nature of light propagation, apodization in high NA imaging systems, and illumination source properties. The derived nonparaxial TCC is numerically compared with the paraxial TCC to demonstrate its added features. The practical forms of the TCC for high resolution phase reconstruction are discussed for two special types of objects, weak objects and slowly varying phase objects. The theoretical studies conducted here can help to bring such high NA microscopy techniques into the real world to solve imaging challenges.
Full-field optical coherence microscopy (FF-OCM) is an imaging technique that provides cross-sectional views of the subsurface microstructure of semitransparent objects. The technology is based on low-coherence interference microscopy, which uses an area camera for en face imaging of the full-field illuminated object. FF-OCM benefits from the lateral imaging resolution of optical microscopy along with the capacity of optical axial sectioning at micrometer-scale resolution. The technique can be employed in diverse applications, in particular for non-invasive examination of biological tissues. This handbook is the first to be entirely devoted to FF-OCM. It is organized into four parts with a total of 21 chapters written by recognized experts and major contributors to the field. After a general introduction to FF-OCM, the fundamental characteristics of the technology are analyzed and discussed theoretically. The main technological developments of FF-OCM for improving the image acquisition speed and for endoscopic imaging are presented in part II. Extensions of FF-OCM for image contrast enhancement or functional imaging are reported in part III. The last part of the book provides an overview of possible applications of FF-OCM in medicine, biology, and materials science. A comprehensive compilation of self-contained chapters written by leading experts, this handbook is a definitive guide to the theoretical analyses, technological developments, and applications of FF-OCM. Using the rich information the book is replete with, a wide range of readers, from scientists and physicists to engineers as well as clinicians and biomedical researchers, can get a handle on the latest major advances in FF-OCM.