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Introduction
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January 2011 - December 2011
September 2012 - present
September 2012 - present
Publications
Publications (239)
Selective Plane Illumination Microscopy (SPIM) has become an emerging technology since its first application for 3D in-vivo imaging of the development of a living organism. An extensive number of works have been published, improving both the speed of acquisition and the resolution of the systems. Furthermore, multispectral imaging allows the effect...
Three-dimensional imaging of live processes at a cellular level is a challenging task. It requires high-speed acquisition capabilities, low phototoxicity, and low mechanical disturbances. Three-dimensional imaging in microfluidic devices poses additional challenges as a deep penetration of the light source is required, along with a stationary setti...
Abnormal cerebral accumulation of amyloid-beta peptide (Aβ) is a major hallmark of Alzheimer’s disease. Non-invasive monitoring of Aβ deposits enables assessing the disease burden in patients and animal models mimicking aspects of the human disease as well as evaluating the efficacy of Aβ-modulating therapies. Previous in vivo assessments of plaque...
Abnormal cerebral accumulation of amyloid-beta peptide (Aβ) is a major hallmark of Alzheimer's disease. Non-invasive monitoring of Aβ deposits enables assessing the disease burden in patients and animal models mimicking aspects of the human disease as well as evaluating the efficacy of Aβ-modulating therapies. Previous in vivo assessments of plaque...
Three-dimensional (3D) optical imaging techniques can expand our knowledge about physiological and pathological processes that cannot be fully understood with 2D approaches. Standard diagnostic tests frequently are not sufficient to unequivocally determine the presence of a pathological condition. Whole organ optical imaging requires tissue transpa...
Developing more efficient methods for antibiotic susceptibility testing is a pressing issue in novel drug development as bacterial resistance to antibiotics becomes increasingly common. Microfluidic devices have been demonstrated to be powerful platforms that allow researchers to perform multiplexed antibiotic testing. However, the level of multipl...
Low scattering media is the best scenario for optical imaging in thick samples and deep tissue, as it allows to obtain high resolution images without suffering the limitations that the diffusion phenomenon imposes. The high contribution of ballistic light in this regime enabled the development of light sheet microscopy and optical projection tomogr...
Light in the near infrared has advantageous properties such as reduced scattering and higher depth of penetration that make it ideal for optical imaging either in transmission or in fluorescence. This chapter gives an overview of the physics behind light propagation, the optical properties of biological tissues and finally presents the potential be...
Fluorescence molecular tomography (FMT) emerges as a powerful non-invasive imaging tool with the ability to resolve fluorescence signals from sources located deep in living tissues. Yet, the accuracy of FMT reconstruction depends on the deviation of the assumed optical properties from the actual values. In this work, we improved the accuracy of the...
Larger than 1 cm sample, cleared with BABB and observed by 405 nm autofluorescence.
SPIM-OPT of 500 angles (0,7º increment/image set)
Fluorescence Molecular Tomography (FMT) constitutes a functional and molecular imaging tool complementing wellestablished anatomical imaging modalities such as magnetic resonance imaging (MRI) and X-ray computer tomography (CT). Preclinical applications of FMT in the fields of tumor imaging, breast imaging, proteomic research, and drug discovery ha...
Objective:
Fluorescence molecular tomography (FMT) can provide valuable molecular information by mapping the bio-distribution of fluorescent reporter molecules in the intact organism. Various prototype FMT systems have been introduced during the past decade. However, none of them has evolved as a standard tool for routine biomedical research. The...
Light-sheet fluorescence microscopy (LSFM) has been present in cell biology laboratories for quite some time, mainly as custom-made systems, with imaging applications ranging from single cells (in the micrometer scale) to small organisms (in the millimeter scale). Such microscopes distinguish themselves for having very low phototoxicity levels and...
Bcl9 and Pygopus (Pygo) are obligate Wnt/β-catenin cofactors in Drosophila, yet their contribution to Wnt signaling during vertebrate development remains unresolved. Combining zebrafish and mouse genetics, we document a conserved, β-catenin-associated function for BCL9 and Pygo proteins during vertebrate heart development. Disrupting the β-catenin-...
LIGATION The animal model of myocardial infarction (MI) plays an important role in the prevention, diagnosis, and therapy of human MI. The study of the behaviour of heart vaso-architecture after MI is produced and its evaluation along time in a 3-dimension (3D) analysis will provide an insight into this disease.
Genetic alterations in human BCL9 genes have repeatedly been found in congenital heart disease (CHD) with as-of-yet unclear causality. BCL9 proteins and their Pygopus (Pygo) co-factors can participate in canonical Wnt signaling via binding to β-catenin. Nonetheless, their contributions to vertebrate heart development remain uncharted. Here, combini...
Optical microscopy constitutes one of the most fundamental paradigms for the understanding of complex biological mechanisms in the whole-organism and live-tissue context. Novel imaging techniques such as light sheet fluorescence microscopy (LSFM) and optical projection tomography (OPT) combined with phase-retrieval algorithms (PRT) can produce high...
We describe a computational method for accurate, quantitative tomographic reconstructions in Optical Projection Tomography, based on phase retrieval algorithms. Our method overcomes limitations imposed by light scattering in opaque tissue samples under the memory effect regime, as well as reduces artifacts due to mechanical movements, misalignments...
3 dimensional (3D) visualization of the localization and distribution of proteins within cells and whole animals is an essential tool to decipher the molecular mechanisms that underlie biological phenomena, such as development, tissue integrity and organismal ageing. The two novel imaging technologies Light Sheet Microscopy (LSM) and Optical Projec...
We present a new Phase-Retrieved Tomography (PRT) method to radically improve mesoscopic imaging at regimes beyond one transport mean-free-path and achieve high resolution, uniformly throughout the volume of opaque samples. The method exploits multi-view acquisition in a hybrid Selective Plane Illumination Microscope (SPIM) and Optical Projection T...
Parenchymal migration of naive CD4(+) T cells in lymph nodes (LNs) is mediated by the Rac activator DOCK2 and PI3Kγ and is widely assumed to facilitate efficient screening of dendritic cells (DCs) presenting peptide-MHCs (pMHCs). Yet how CD4(+) T cell motility, DC density, and pMHC levels interdependently regulate such interactions has not been com...
The ability to acquire 3D images of the heart and its vasculature at cellular resolution facilitates a more detailed study of many heart diseases. Here, we describe a novel technique to image in 3D the heart vasculature by combining the CUBIC clearing protocol combined with in vivo administration of fluorescent-labeled lectin. The use of these tech...
The CUBIC tissue clearing protocol has been optimized to produce translucent immunostained whole chicken embryos and embryo brains. When combined with multispectral light sheet microscopy, the validated protocol presented here provides a rapid, inexpensive and reliable method for acquiring accurate histological images that preserve three-dimensiona...
Light sheet fluorescence microscopy enables high resolution imaging of thick biological samples. By restricting the fluorescence excitation to a single plane, rapid wide-field image acquisition is possible with minimal sample exposure. Although light sheet microscopy is able to resolve sub-cellular features at depth in model organisms, elevated lev...
Diffuse Optical Tomography commonly neglects or assumes as insignificant the presence of optically clear regions in biological tissues, estimating their contribution as a small pertur-bation to light transport. The inaccuracy introduced by this prac-tice is examined in detail in the context of a complete, based on realistic geometry, virtual fluore...
During adaptive immune responses, CD8(+) T cells with low TCR affinities are released early into the circulation before high-affinity clones become dominant at later time points. How functional avidity maturation is orchestrated in lymphoid tissue and how low-affinity cells contribute to host protection remains unclear. In this study, we used intra...
Stripe artifacts, caused by high-absorption or high-scattering structures in the illumination light path, are a common drawback in both unidirectional and multidirectional light sheet fluorescence microscopy (LSFM), significantly deteriorating image quality. To circumvent this problem, we present an effective multidirectional stripe remover (MDSR)...
The ability to acquire high resolution 3D images of the heart enables to study heart diseases more in detail. In this work, the CUBIC (clear, unobstructed brain imaging cocktails and computational analysis) clearing protocol was optimized for thick mouse heart sections to enhance the penetration depth of the confocal microscope lasers into the tiss...
Multispectral tissue imaging based on optical cameras and continuous-wave tissue illumination is commonly used in medicine and biology. Surprisingly, there is a characteristic absence of a critical look at the quantities that can be uniquely characterized from optically diffuse matter by multispectral imaging. Here, we investigate the fundamental q...
Optical Neuroimaging is a highly dynamical field of research owing to the combination of many advanced imaging techniques and computational tools that uncovered unexplored paths through the functioning of the brain. Light propagation modelling through such complicated structures has always played a crucial role as the basis for a high resolution an...
Of the large amount of the animal models available for cardiac research, the zebrafish is extremely valuable due to its transparency during early stages of development. In this work a dual illumination laser sheet microscope with simultaneous dual camera imaging is used to image the beating heart at 200 fps, dynamically and selectively focusing ins...
We here show 3D light sheet microscopy images of fixed and cleared murine colon tissue in-toto, which offer relevant cellular information without the need for physically sectioning the tissue. We have applied the recently developed CUBIC protocol (Susaki et al. Cell 157:726, 2014) for colon tissues and have found that this clearing protocol enables...
Fluorescent proteins and dyes are routine tools for biological research to describe the behavior of genes, proteins, and cells, as well as more complex physiological dynamics such as vessel permeability and pharmacokinetics. The use of these probes in whole body in vivo imaging would allow extending the range and scope of current biomedical applica...
Reactive lymph nodes (LNs) are sites where pMHC-loaded dendritic cells (DCs) interact with rare cognate T cells, leading to their clonal expansion. While DC interactions with T cell subsets critically shape the ensuing immune response, surprisingly little is known on their spatial orchestration at physiologically T cell low precursor frequencies. L...
Optical projection tomography (OPT) is a tool used for three-dimensional imaging of millimeter-scale biological samples, with the advantage of exhibiting isotropic resolution typically in the micron range. OPT can be divided into two types: transmission OPT (tOPT) and emission OPT (eOPT). Compared with eOPT, tOPT discriminates different tissues bas...
In this talk we will show that the physical measurements of light emerging from a diffusive medium may be used directly to solve the inverse problem, producing more accurate results due to the fact that the actual surface profile is used.
Imaging approaches are an essential tool for following up over time representative parameters of in vivo models, providing useful information in pharmacological studies. Main advantages of optical imaging approaches compared to other imaging methods are their safety, straight-forward use and cost-effectiveness. A main drawback, however, is having t...
The past decade marked an optical revolution in biology: an unprecedented number of optical techniques were developed and adopted for biological exploration, demonstrating increasing interest in optical imaging and in vivo interrogations. Optical methods have become faster and have reached nanoscale resolution, and are now complemented by optoacous...
Purpose:
Assessing tumor vascular features including permeability and perfusion is essential for diagnostic and therapeutic purposes. The aim of this study was to compare fluorescence and magnetic resonance imaging (MRI)-based vascular readouts in subcutaneously implanted tumors in mice by simultaneous dynamic measurement of tracer uptake using a...
One of the major challenges within Optical Imaging, photon propagation through clear layers embedded between scattering tissues, can be now efficiently modelled in real-time thanks to the Monte Carlo approach based on GPU. Because of its nature, the photon propagation problem can be very easily parallelized and ran on low cost hardware, avoiding th...
Endoscopy is frequently used in the diagnosis of several gastro-intestinal pathologies as Crohn disease, ulcerative colitis or colorectal cancer. It has great potential as a non-invasive screening technique capable of detecting suspicious alterations in the intestinal mucosa, such as inflammatory processes. However, these early lesions usually cann...
We describe a customizable and cost-effective light sheet microscopy (LSM) platform for rapid three-dimensional imaging of protein dynamics in small model organisms. The system is designed for high acquisition speeds and enables extended time-lapse in vivo experiments when using fluorescently labeled specimens. We demonstrate the capability of the...
Introduction
Fluorescence Molecular Tomography (FMT) suffers from poor spatial resolution and lack of anatomy information, which may be accounted for by combining the method with high resolution (HR) imaging modalities such as X-ray CT and MRI. We have a non-contact FMT/MRI hybrid system [1]. In this study we explored the potential of this hybrid s...
Optical Projection Tomography (OPT) is a 3-Dimentional (3D) imaging technique for small specimens between 1mm and 10mm in size. Due to its high resolution and whole-body imaging ability, OPT has been widely used for imaging of small specimens such as murine embryos, murine organs, zebra fish, and plant sections. During an OPT imaging experiment, th...
A method for tomographic imaging of diffuse medium includes directing waves into a diffusive medium, solving a surface-bounded inversion problem by forward field calculations through decomposition of contributions from the multiple reflections from an arbitrary surface within the diffusive medium or outside the diffusive medium into a sum of differ...
Optical imaging technologies such as fluorescence molecular tomography (FMT) are gaining great relevance in cardiovascular research. The main reason is the increased number of available fluorescent agents, especially those termed “activatable probes,” which remain quenched under baseline conditions and are fluorescent when a specific enzymatic acti...
Even though in vivo imaging approaches have witnessed several new and important developments, specimens that exhibit high light scattering properties such as Drosophila melanogaster pupae are still not easily accessible with current optical imaging techniques, obtaining images only from subsurface features. This means that in order to obtain 3D vol...
Optical Projection Tomography and Light Sheet Microscopy have been recent revolutionary techniques for imaging in biological specimens with 3D volume rendering of high resolution. A comparison study is performed on the basic aspects of each modality and combined application on selected specimen is presented. The imaging performance characteristics...
Optical Projection Tomography and Light Sheet Microscopy have been recent revolutionary techniques for imaging in biological specimens with 3D volume rendering of high resolution. A comparison study is performed on the basic aspects of each modality and combined application on selected specimen is presented. The imaging performance characteristics...
Laser sheet microscopy is a widely used imaging technique for imaging the three-dimensional distribution of a fluorescence signal in fixed tissue or small organisms. In laser sheet microscopy, the stripe artifacts caused by high absorption or high scattering structures are very common, greatly affecting image quality. To solve this problem, we repo...
A new method to obtain the three-dimensional localization of fluorochrome distributions in micrometric samples is presented. It uses a microlens array coupled to the image port of a standard microscope to obtain tomographic data by a filtered back-projection algorithm. Scanning of the microlens array is proposed to obtain a dense data set for recon...
The increasing preclinical and clinical utilization of digital cameras for photographic measurements of tissue conditions motivates the study of reflectance measurements obtained with planar illumination. We examine herein a formula that models the total diffuse reflectance measured from a semi-infinite medium using an exponentially decaying source...
Compressed Sensing (CS) techniques are becoming increasingly popular to speed up data acquisition in many modalities. However, most of CS theory is devoted to undetermined problems and there are few contributions that apply it to ill-posed problems. In this work we present a novel approach to CS for fluorescence diffuse optical tomography (fDOT), n...
A new technique termed Helical Optical Projection Tomography (hOPT) has been developed with the aim to overcome some of the limitations of current 3D optical imaging techniques. hOPT is based on Optical Projection Tomography (OPT) with the major difference that there is a translation of the sample in the vertical direction during the image acquisit...
Modeling light propagation in the whole body is essential and necessary for optical imaging. However, non-scattering, low-scattering and high absorption regions commonly exist in biological tissues, which lead to inaccuracy of the existing light transport models. In this paper, a novel hybrid light transport model that couples the simplified spheri...
Recent advances in biomedical imaging technologies have revolutionized the way we approach a variety of medical and biological questions and have unveiled several biological processes, mechanisms and functions. Optical Imaging methods facilitate the use of light for the non-invasive visualization of structure and function of biological tissue with...
Even though endoscopic fluorescence diffuse optical tomography (eFDOT) exhibits significant potential, currently its application is limited due to the existence of strong autofluorescence and the imaging inaccuracy caused by a very short source-detector distance. Motivated by the emerging X-ray luminescence computed tomography (XLCT) technology, we...
Fluorescence diffuse optical tomography (fDOT) is a noninvasive imaging technique that makes it possible to quantify the spatial distribution of fluorescent tracers in small animals. fDOT image reconstruction is commonly performed by means of iterative methods such as the algebraic reconstruction technique (ART). The useful results yielded by more...
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In Optical Projection Tomography (OPT), if the rotational center deviates from the central line of the image and this offset is not corrected during the reconstruction, serious blurring will happen in the final 3-dimensional (3D) result. Therefore, the high-precision rotational center location method is very important for OPT. However, existing met...
During the last decades, especially via the EU initiative related to the Virtual Physiological Human, significant progress has been made in advancing "in-silico" computational models to produce accurate and reliable tumor growth simulations. However, currently most attempts to validate the outcome of the models are either done in-vitro or ex-vivo a...
We present a custom-made combined, selective plane illumination microscopy (SPIM) and optical projection tomography (OPT) system for rapid three-dimensional imaging. OPT eliminates the need for serial optical sectioning of specimens, required with conventional confocal microscopy, and allows for robust, high resolution fluorescence, as well as, abs...
The invention relates to systems and methods for tomographic imaging in diffuse media employing a fast reconstruction technique. A hybrid Fourier approach is presented that enables the fast tomographic reconstruction of large datasets. In certain embodiments, the invention features methods of in vivo fluorescence molecular tomographic (FMT) reconst...
The invention relates to systems and methods for tomographic imaging of a subject comprising diffuse media by converting measurements of electromagnetic radiation, e.g., fluorescent light, obtained in free space exterior to the subject into data that would be measured if the subject were surrounded by an infinite and homogeneous diffusive medium, e...