In Vivo Tomographic Imaging of Near-infrared Fluorescent Probes

Molecular Imaging. Apr-Jun, 2002  |  Pubmed ID: 12920848

Fluorescence imaging is increasingly used to probe protein function and gene expression in live animals. This technology could enhance the study of pathogenesis, drug development, and therapeutic intervention. In this article, we focus on three-dimensional fluorescence observations using fluorescence-mediated molecular tomography (FMT), a novel imaging technique that can resolve molecular function in deep tissues by reconstructing fluorescent probe distributions in vivo. We have compared FMT findings with conventional fluorescence reflectance imaging (FRI) to study protease function in nude mice with subsurface implanted tumors. This validation of FMT with FRI demonstrated the spatial congruence of fluorochrome activation as determined by the two techniques.

High Throughput Magnetic Resonance Imaging for Evaluating Targeted Nanoparticle Probes

Bioconjugate Chemistry. Jan-Feb, 2002  |  Pubmed ID: 11792186

The ability to image specific molecular targets in vivo would have significant impact in allowing earlier disease detection and in tailoring molecular therapies. One of the rate-limiting steps in the development of novel compounds as reporter probes has been the lack of cell-based, biologically relevant, high throughput screening methods. Here we describe the development and validation of magnetic resonance imaging (MRI) as a technique to rapidly screen compounds that are potential MR reporter agents for their interaction with specific cellular targets. We show that MR imaging can (1) evaluate thousands of samples simultaneously and rapidly, (2) provide exceedingly accurate measurements, and (3) provide receptor binding/internalization data as validated by radioactive assays. The technique allows the screening of libraries of peptide-nanoparticle conjugates against target cells and the identification of conjugates that may be subsequently used as reporter agents in vivo. The technology should greatly accelerate the development of target-specific or cell-specific MR contrast agents.

Projection Access Order in Algebraic Reconstruction Technique for Diffuse Optical Tomography

Physics in Medicine and Biology. Jan, 2002  |  Pubmed ID: 11814231

Algebraic reconstruction technique (ART) is one of the popular image reconstruction techniques used in diffuse optical tomography (DOT). We investigate in this note the influence of the order in which data are accessed in ART. Simulations mimicking breast tissues in transmission geometry with contrast agent tumour enhancement were used to evaluate the image quality of the diverse projection access investigated. We show that by selecting proper projection access order, the convergence speed can be significantly improved when ART is used to perform DOT. Moreover, low-contrast detection is improved.

Charge-coupled-device Based Scanner for Tomography of Fluorescent Near-infrared Probes in Turbid Media

Medical Physics. May, 2002  |  Pubmed ID: 12033576

We present a novel tomographer for three-dimensional reconstructions of fluorochromes in diffuse media. Photon detection is based on charge-coupled device technology that allows the implementation of a large parallel array of detection channels with high sensitivity. Using this instrument we studied the response and detection limits of near-infrared fluorochromes in diffuse media as a function of light intensity and for a wide range of biologically relevant concentrations. We further examined the resolution of the scanner and the reconstruction linearity achieved. We demonstrate that the instrument attains better than 3 mm resolution, is linear within more than two orders of magnitude of fluorochrome concentration, and can detect fluorescent objects at femto-mole quantities in small animal-like geometries. These measurements delineate detection and reconstruction characteristics associated with imaging of novel classes of fluorescent probes developed for in vivo molecular and functional probing of tissues.

In Vivo Imaging of Proteolytic Activity in Atherosclerosis

Circulation. Jun, 2002  |  Pubmed ID: 12057992

Atherosclerotic plaque rupture, the most important cause of acute cardiovascular incidents, has been strongly associated with vascular inflammation. On the basis of the hypothesis that the inflammatory response and proteolysis lead to plaque rupture, we have examined the role of cathepsin B as a model proteolytic enzyme.

MRI-guided Diffuse Optical Spectroscopy of Malignant and Benign Breast Lesions

Neoplasia (New York, N.Y.). Jul-Aug, 2002  |  Pubmed ID: 12082551

We present the clinical implementation of a novel hybrid system that combines magnetic resonance imaging (MRI) and near-infrared (NIR) optical measurements for the noninvasive study of breast cancer in vivo. Fourteen patients were studied with a MR-NIR prototype imager and spectrometer. A diffuse optical tomographic scheme employed the MR images as a priori information to implement an image-guided NIR localized spectroscopic scheme. All patients who entered the study also underwent gadolinium-enhanced MRI and biopsy so that the optical findings were cross-validated with MR readings and histopathology. The technique quantified the oxy- and deoxyhemoglobin of five malignant and nine benign breast lesions in vivo. Breast cancers were found with decreased oxygen saturation and higher blood concentration than most benign lesions. The average hemoglobin concentration ([H]) of cancers was 0.130+/-0.100 mM, and the average hemoglobin saturation (Y) was 60+/-9% compared to [H]=0.018+/-0.005 mM and Y=69+/-6% of background tissue. Fibroadenomas exhibited high hemoglobin concentration [H]=0.060+/-0.010 mM and mild decrease in oxygen saturation Y=67+/-2%. Cysts and other normal lesions were easily differentiated based on intrinsic contrast information. This novel optical technology can be a significant add-on in MR examinations and can be used to characterize functional parameters of cancers with diagnostic and treatment prognosis potential. It is foreseen that the technique can play a major role in functional activation studies of brain and muscle as well.

Fluorescence Molecular Tomography Resolves Protease Activity in Vivo

Nature Medicine. Jul, 2002  |  Pubmed ID: 12091907

Systematic efforts are under way to develop novel technologies that would allow molecular sensing in intact organisms in vivo. Using near-infrared fluorescent molecular beacons and inversion techniques that take into account the diffuse nature of photon propagation in tissue, we were able to obtain three-dimensional in vivo images of a protease in orthopic gliomas. We demonstrate that enzyme-activatable fluorochromes can be detected with high positional accuracy in deep tissues, that molecular specificities of different beacons towards enzymes can be resolved and that tomography of beacon activation is linearly related to enzyme concentration. The tomographic imaging method offers a range of new capabilities for studying biological function; for example, identifying molecular-expression patterns by multispectral imaging or continuously monitoring the efficacy of therapeutic drugs.

Imaging Cathepsin B Up-regulation in HT-1080 Tumor Models Using Fluorescence-mediated Molecular Tomography (FMT)

Academic Radiology. Aug, 2002  |  Pubmed ID: 12188262

Would Near-infrared Fluorescence Signals Propagate Through Large Human Organs for Clinical Studies?

Optics Letters. Mar, 2002  |  Pubmed ID: 18007794

We predict the capacity of near-infrared fluorescent signals to propagate through human tissue for non-invasive medical imaging. This analysis employs experimental measurements of a biologically relevant local fluorochrome embedded in tissuelike media and predicts the equivalent photon counts expected from breast, lung, brain, and muscle as a function of diameter by use of an analytical solution of the diffusion equation that can take into account large arbitrary geometries. The findings address feasibility issues for clinical studies and are relevant to recent development of near-infrared fluorescent probes and molecular beacons for in vivo applications.

Fast Analytical Approximation for Arbitrary Geometries in Diffuse Optical Tomography

Optics Letters. Apr, 2002  |  Pubmed ID: 18007854

Diffuse optical tomography is a novel imaging technique that resolves and quantifies the optical properties of objects buried in turbid media. Typically, numerical solutions of the diffusion equation are employed to construct the tomographic problem when media of complex geometries are investigated. Numerical methods offer implementation simplicity but also significant computation burden, especially when large three-dimensional reconstructions are involved. We present an alternative method of performing tomography of diffuse media of arbitrary geometries by means of an analytical approach, the Kirchhoff approximation. We show that the method is extremely efficient in computation times and consider its potential as a real-time three-dimensional imaging tool.

Would Near-infrared Fluorescence Signals Propagate Through Large Human Organs for Clinical Studies? Errata

Optics Letters. Sep, 2002  |  Pubmed ID: 18026531

Analytical Model for Dual-interfering Sources Diffuse Optical Tomography

Optics Express. Jan, 2002  |  Pubmed ID: 19424324

An analytical model to perform tomographic reconstructions for absorptive inclusions in highly scattering media using dual interfering sources was derived. A perturbation approach within the first order Rytov expansion was used to solve the heterogeneous diffusion equation. Analytical weight functions necessary to solve the inverse problem were obtained. The reconstructions performance was assessed using simulated data of breast-like media after contrast agent enhancement. We further investigated the reconstruction quality as a function of object depth location, modulation frequency and source separation. The ability of the algorithm to resolve multi-objects was also demonstrated.

Steady-state Blood Volume Measurements in Experimental Tumors with Different Angiogenic Burdens a Study in Mice

Radiology. Jan, 2003  |  Pubmed ID: 12511693

To experimentally validate the effectiveness of magnetic resonance (MR) imaging enhanced with long-circulating iron oxide for measurement of vascular volume fractions (VVFs) as indicators of angiogenesis in different experimental tumor models.

Shedding Light Onto Live Molecular Targets

Nature Medicine. Jan, 2003  |  Pubmed ID: 12514725

Fluorescence Imaging with Near-infrared Light: New Technological Advances That Enable in Vivo Molecular Imaging

European Radiology. Jan, 2003  |  Pubmed ID: 12541130

A recent development in biomedical imaging is the non-invasive mapping of molecular events in intact tissues using fluorescence. Underpinning to this development is the discovery of bio-compatible, specific fluorescent probes and proteins and the development of highly sensitive imaging technologies for in vivo fluorescent detection. Of particular interest are fluorochromes that emit in the near infrared (NIR), a spectral window, whereas hemoglobin and water absorb minimally so as to allow photons to penetrate for several centimetres in tissue. In this review article we concentrate on optical imaging technologies used for non-invasive imaging of the distribution of such probes. We illuminate the advantages and limitations of simple photographic methods and turn our attention to fluorescence-mediated molecular tomography (FMT), a technique that can three-dimensionally image gene expression by resolving fluorescence activation in deep tissues. We describe theoretical specifics, and we provide insight into its in vivo capacity and the sensitivity achieved. Finally, we discuss its clinical feasibility.

Probing Rat Brain Oxygenation with Near-infrared Spectroscopy (NIRS) and Magnetic Resonance Imaging (MRI)

Advances in Experimental Medicine and Biology. 2003  |  Pubmed ID: 12580428

Optical-based Molecular Imaging: Contrast Agents and Potential Medical Applications

European Radiology. Feb, 2003  |  Pubmed ID: 12598985

Laser- and sensitive charge-coupled device technology together with advanced mathematical modelling of photon propagation in tissue has prompted the development of novel optical imaging technologies. Fast surface-weighted imaging modalities, such as fluorescence reflectance imaging (FRI) and 3D quantitative fluorescence-mediated tomography have now become available [1, 2]. These technical advances are paralleled by a rapid development of a whole range of new optical contrasting strategies, which are designed to generate molecular contrast within a living organism. The combination of both, technical advances of light detection and the refinement of optical contrast media, finally yields a new spectrum of tools for in vivo molecular diagnostics. Whereas the technical aspects of optical imaging are covered in more detail in a previous review article in "European Radiology" [3], this article focuses on new developments in optical contrasting strategies and design of optical contrast agents for in vivo diagnostics.

A Submillimeter Resolution Fluorescence Molecular Imaging System for Small Animal Imaging

Medical Physics. May, 2003  |  Pubmed ID: 12772999

Most current imaging systems developed for tomographic investigations of intact tissues using diffuse photons suffer from a limited number of sources and detectors. In this paper we describe the construction and evaluation of a large dataset, low noise tomographic system for fluorescence imaging in small animals. The system consists of a parallel plate-imaging chamber and a lens coupled CCD camera, which enables conventional planar imaging as well as fluorescence tomography. The planar imaging data are used to guide the acquisition of a Fluorescence Molecular Tomography (FMT) dataset containing more than 106 measurements, and to superimpose anatomical features with tomographic results for improved visual representation. Experimental measurements exhibited good agreement with the diffusion theory models used to predict light propagation within the chamber. Tests of the instrument's capacity to quantitatively reconstruct fluorochrome distributions in three dimensions showed less than 5% errors between actual fluorochrome concentrations and FMT findings, and suggested a detection threshold of approximately 100 femptomoles for small localized objects. Experiments to assess the instrument's spatial resolution demonstrated the ability of the system to resolve objects placed at clear distances of less than 1 mm. This is a significant resolution increase over previously developed systems for animal imaging, and is primarily due to the large dataset employed and the use of inversion methods. Finally, the in vivo imaging capacity is showcased. It is expected that the large dataset collected can enable superior imaging of molecular probes in vivo and improve quantification of fluorescence signatures.

Iterative Boundary Method for Diffuse Optical Tomography

Journal of the Optical Society of America. A, Optics, Image Science, and Vision. Jun, 2003  |  Pubmed ID: 12801178

The recent application of tomographic methods to three-dimensional imaging through tissue by use of light often requires modeling of geometrically complex diffuse-nondiffuse boundaries at the tissue-air interface. We have recently investigated analytical methods to model complex boundaries by means of the Kirchhoff approximation. We generalize this approach using an analytical approximation, the N-order diffuse-reflection boundary method, which considers higher orders of interaction between surface elements in an iterative manner. We present the general performance of the method and demonstrate that it can improve the accuracy in modeling complex boundaries compared with the Kirchhoff approximation in the cases of small diffuse volumes or low absorption. Our observations are also contrasted with exact solutions. We furthermore investigate optimal implementation parameters and show that a second-order approximation is appropriate for most in vivo investigations.

Optical Imaging of Apoptosis As a Biomarker of Tumor Response to Chemotherapy

Neoplasia (New York, N.Y.). May-Jun, 2003  |  Pubmed ID: 12869301

A rapid and accurate assessment of the antitumor efficacy of new therapeutic drugs could speed up drug discovery and improve clinical decision making. Based on the hypothesis that most effective antitumor agents induce apoptosis, we developed a near-infrared fluorescent (NIRF) annexin V to be used for optical sensing of tumor environments. To demonstrate probe specificity, we developed both an active (i.e., apoptosis-recognizing) and an inactive form of annexin V with very similar properties (to account for nonspecific tumor accumulation), and tested the agents in nude mice each bearing a cyclophosphamide (CPA) chemosensitive (LLC) and a chemoresistant LLC (CR-LLC). After injection with active annexin V, the tumor-annexin V ratio (TAR; tumor NIRF/background NIRF) for untreated mice was 1.22+/-0.34 for LLC and 1.43+/-0.53 for CR-LLC (n=4). The LLC of CPA-treated mice had significant elevations of TAR (2.56+/-0.29, P=.001, n=4), but only a moderate increase was obtained for the CR-LLC (TAR=1.89+/-0.19, P=.183). The in vivo measurements correlated well with terminal deoxyribosyl transferase-mediated dUTP nick end labeling indexes. When inactive Cy-annexin V was used, with or without CPA treatment and in both CCL and CR-CCL tumors, tumor NIRF values ranged from 0.91 to 1.17 (i.e., tumor were equal to background). We conclude that active Cy-annexin V and surface reflectance fluorescence imaging provide a nonradioactive, semiquantitative method of determining chemosensitivity in LLC xenografts. The method maybe used to image pharmacologic responses in other animal models and, potentially, may permit the clinical imaging of apoptosis with noninvasive or minimally invasive instrumentation.

Noncontact Optical Tomography of Turbid Media

Optics Letters. Sep, 2003  |  Pubmed ID: 13677542

Optical tomography of turbid media has so far been limited by systems that require fixed geometries or measurements employing fibers. We present a system that records noncontact optical measurements from diffuse media of arbitrary shapes and retrieves the three-dimensional surface information of the diffuse medium. We further present a novel method of combining this composite data set and obtain accurate fluorescence reconstructions. This approach offers significant experimental simplicity and yields high-information-content datasets. The performance of this novel tomographic approach is demonstrated with experimental reconstructions of phantoms.

Free-space Propagation of Diffuse Light: Theory and Experiments

Physical Review Letters. Sep, 2003  |  Pubmed ID: 14525478

In this Letter we present a simple and novel theoretical approach for modeling the intensity distribution from an arbitrarily shaped turbid volume in a noncontact geometry by considering diffuse light propagation in free space. This theory is validated with experiments for a diffusive volume of known geometry in a noncontact situation, both with and without the presence of an embedded absorber. The implications of this new formulation in the context of optical tomography in turbid media are discussed.

Singular-value Analysis and Optimization of Experimental Parameters in Fluorescence Molecular Tomography

Journal of the Optical Society of America. A, Optics, Image Science, and Vision. Feb, 2004  |  Pubmed ID: 14763766

The advent of specific molecular markers and probes employing optical reporters has encouraged the application of in vivo diffuse tomographic imaging at greater spatial resolutions and hence data-set volumes. This study applied singular-value analysis (SVA) of the fluorescence tomographic problem to determine optimal source and detector distributions that result in data sets that are balanced between information content and size. Weight matrices describing the tomographic forward problem were constructed for a range of source and detector distributions and fields of view and were decomposed into their associated singular values. These singular-value spectra were then compared so that we could observe the effects of each parameter on imaging performance. The findings of the SVA were then confirmed by examining reconstructions of simulated and experimental data acquired with the same optode distributions as examined by SVA. It was seen that for a 20-mm target width, which is relevant to the small-animal imaging situation, the source and detector fields of view should be set at approximately 30 mm. Equal numbers of sources and detectors result in the best imaging performance in the parallel-plate geometry and should be employed when logistically feasible. These data provide guidelines for the design of small-animal diffuse optical tomographic imaging systems and demonstrate the utility of SVA as a simple and efficient means of optimizing experimental parameters in problems for which a forward model of the data collection process is available.

Experimental Fluorescence Tomography of Tissues with Noncontact Measurements

IEEE Transactions on Medical Imaging. Apr, 2004  |  Pubmed ID: 15084074

Noncontact optical measurements from diffuse media could facilitate the use of large detector arrays at multiple angles that are well suited for diffuse optical tomography applications. Such imaging strategy could eliminate the need for individual fibers in contact with tissue, restricted geometries, and matching fluids. Thus, it could significantly improve experimental procedures and enhance our ability to visualize functional and molecular processes in vivo. In this paper, we describe the experimental implementation of this novel concept and demonstrate capacity to perform small animal imaging.

Visualization of Antitumor Treatment by Means of Fluorescence Molecular Tomography with an Annexin V-Cy5.5 Conjugate

Proceedings of the National Academy of Sciences of the United States of America. Aug, 2004  |  Pubmed ID: 15304657

In vivo imaging of treatment responses at the molecular level could have a significant impact on the speed of drug discovery and ultimately lead to personalized medicine. Strong interest has been shown in developing quantitative fluorescence-based technologies with good molecular specificity and sensitivity for noninvasive 3D imaging through tissues and whole animals. We show herein that tumor response to chemotherapy can be accurately resolved by fluorescence molecular tomography (FMT) with a phosphatidylserine-sensing fluorescent probe based on modified annexins. We observed at least a 10-fold increase of fluorochrome concentration in cyclophosphamide-sensitive tumors and a 7-fold increase of resistant tumors compared with control studies. FMT is an optical imaging technique developed to overcome limitations of commonly used planar illumination methods and demonstrates higher quantification accuracy validated by histology. It is further shown that a 3-fold variation in background absorption heterogeneity may yield 100% errors in planar imaging but only 20% error in FMT, thus confirming tomographic imaging as a preferred tool for quantitative investigations of fluorescent probes in tissues. Tomographic approaches are found essential for small-animal optical imaging and are potentially well suited for clinical drug development and monitoring.

Complete-angle Projection Diffuse Optical Tomography by Use of Early Photons

Optics Letters. Feb, 2005  |  Pubmed ID: 15762444

We present the first, to our knowledge, experimental images of complex-shaped phantoms embedded in diffuse media by use of optical tomography. Imaging is based on a complete-angle projection tomographic technique that utilizes transmitted early photons. Results are contrasted with measurements obtained at later gates as well as pseudocontinuous-wave data. The scanning system developed employs noncontact illumination and detection technologies that allow for high spatial sampling of transmitted photons. Combining this system with complete-angle illumination is found to be an important strategy toward improved imaging performance, resulting in a better-posed inversion problem. The appropriateness of reconstruction algorithms similar to those employed in x-ray computed tomography are showcased, and suggestions for model improvements are provided.

Looking and Listening to Light: the Evolution of Whole-body Photonic Imaging

Nature Biotechnology. Mar, 2005  |  Pubmed ID: 15765087

Optical imaging of live animals has grown into an important tool in biomedical research as advances in photonic technology and reporter strategies have led to widespread exploration of biological processes in vivo. Although much attention has been paid to microscopy, macroscopic imaging has allowed small-animal imaging with larger fields of view (from several millimeters to several centimeters depending on implementation). Photographic methods have been the mainstay for fluorescence and bioluminescence macroscopy in whole animals, but emphasis is shifting to photonic methods that use tomographic principles to noninvasively image optical contrast at depths of several millimeters to centimeters with high sensitivity and sub-millimeter to millimeter resolution. Recent theoretical and instrumentation advances allow the use of large data sets and multiple projections and offer practical systems for quantitative, three-dimensional whole-body images. For photonic imaging to fully realize its potential, however, further progress will be needed in refining optical inversion methods and data acquisition techniques.

Experimental Determination of Photon Propagation in Highly Absorbing and Scattering Media

Journal of the Optical Society of America. A, Optics, Image Science, and Vision. Mar, 2005  |  Pubmed ID: 15770993

Optical imaging and tomography in tissues can facilitate the quantitative study of several important chromophores and fluorophores. Several theoretical models have been validated for diffuse photon propagation in highly scattering and low-absorbing media that describe the optical appearance of tissues in the near-infrared (NIR) region. However, these models are not generally applicable to quantitative optical investigations in the visible because of the significantly higher tissue absorption in this spectral region compared with that in the NIR. We performed photon measurements through highly scattering and absorbing media for ratios of the absorption coefficient to the reduced scattering coefficient ranging approximately from zero to one. We examined experimentally the performance of the absorption-dependent diffusion coefficient defined by Aronson and Corngold [J. Opt. Soc. Am. A 16, 1066 (1999)] for quantitative estimations of photon propagation in the low- and high-absorption regimes. Through steady-state measurements we verified that the transmitted intensity is well described by the diffusion equation by considering a modified diffusion coefficient with a nonlinear dependence on the absorption. This study confirms that simple analytical solutions based on the diffusion approximation are suitable even for high-absorption regimes and shows that diffusion-approximation-based models are valid for quantitative measurements and tomographic imaging of tissues in the visible.

Quantitative Point Source Photoacoustic Inversion Formulas for Scattering and Absorbing Media

Physical Review. E, Statistical, Nonlinear, and Soft Matter Physics. Mar, 2005  |  Pubmed ID: 15903464

We present here an expression for the photoacoustic contribution of an optical point source in a diffusive and absorbing medium. By using this measurement as a reference, we present a direct inversion formula that recovers the absorption map quantitatively, at the same time accounting for instrumental factors such as the source strength, the shape of the optical pulse, and the impulse response and finite size of the transducers. We further validate this expression through accurate numerical simulations showing that the absorption map is recovered quantitatively in the presence of a rotating geometry. We finally discuss how the presented solutions for point sources within the photoacoustic problem enable the use of concurrent fluorescence and ultrasound measurements as appropriate for a hybrid tomographic system. The proposed system could retrieve absorption information using photoacoustic measurements, and use these data to more accurately describe the fluorescence problem and improve reconstruction fidelity.

Optical Imaging of Spontaneous Breast Tumors Using Protease Sensing 'smart' Optical Probes

Investigative Radiology. Jun, 2005  |  Pubmed ID: 15905717

The objective of this study was to determine if spontaneous breast cancer lesions can be detected by fluorescence reflectance imaging (FRI) and fluorescence mediated tomography (FMT) using protease-sensing optical probes.

Fluorescent Protein Tomography Scanner for Small Animal Imaging

IEEE Transactions on Medical Imaging. Jul, 2005  |  Pubmed ID: 16011317

Microscopy of fluorescent proteins has enabled unprecedented insights into visualizing gene expression in living systems. Imaging deeper into animals, however, has been limited due to the lack of accurate imaging methods for the visible. We present a novel system designed to perform tomographic imaging of fluorescent proteins through whole animals. The tomographic method employed a multiangle, multiprojection illumination scheme, while detection was achieved using a highly sensitive charge-coupled device camera with appropriate filters. Light propagation was modeled using a modified solution to the diffusion equation to account for the high absorption and high scattering of tissue at the visible wavelengths. We show that the technique can quantitatively detect fluorescence with sub millimeter spatial resolution both in phantoms and in tissues. We conclude that the method could be applied in tomographic imaging of fluorescent proteins for in vivo targeting of different diseases and abnormalities.

Image Analysis for Assessing Molecular Activity Changes in Time-dependent Geometries

IEEE Transactions on Medical Imaging. Jul, 2005  |  Pubmed ID: 16011319

In vivo fluorescence molecular imaging and tomography has facilitated monitoring of genomics and proteomics over time and on the same animal. A highly important issue, however, has been the robust registration of animals imaged at different time points to obtain accurate description of activity and location. This paper presents a method for aligning temporal data of small animals based on surface anatomical features and improving the accuracy of monitoring fluorophore distribution. The method can account for differences in the positioning and compression of small animals and can be extended to three-dimensional as well as to other imaging modalities.

Tomographic Fluorescence Mapping of Tumor Targets

Cancer Research. Jul, 2005  |  Pubmed ID: 16024635

Methods that allow robust imaging of specific molecular targets and biological processes in vivo should have widespread applications in biology and clinical medicine. Here we use a quantitative, three-dimensional fluorescence-mediated tomographic technique (FMT) that enables rapid measurements of fluorochrome-based affinity tags in live xenograft models. We validate the method by showing its sensitivity in quantitating tumor angiogenesis and therapeutic modulation using an anti-vascular endothelial growth factor antibody. Furthermore, we show the feasibility of simultaneous multichannel measurements of distinct biological phenomena such as receptor tyrosine kinase expression and angiogenesis. FMT measurements can be done serially, with short imaging times and within the same live animal. The described method should be valuable for rapidly profiling biological phenomena in vivo.

Quantitative Analysis of Chemotherapeutic Effects in Tumors Using in Vivo Staining and Correlative Histology

Cellular Oncology : the Official Journal of the International Society for Cellular Oncology. 2005  |  Pubmed ID: 16037639

To microscopically analyze the chemotherapeutic response of tumors using in vivo staining based on an annexinV-Cy5.5 probe and independently asses their apoptotic count using quantitative histological analysis.

Effects of Background Fluorescence in Fluorescence Molecular Tomography

Applied Optics. Sep, 2005  |  Pubmed ID: 16161661

Recent advances in optical imaging systems and systemically administered fluorescent probes have significantly improved the ways by which we can visualize proteomics in vivo. A key component in the design of fluorescent probes is a favorable biodistribution, i.e., localization only in the targeted diseased tissue, in order to achieve high contrast and good detection characteristics. In practice, however, there is always some level of background fluorescence present that could result in distorted or obscured visualization and quantification of measured signals. In this study we observe the effects of background fluorescence in tomographic imaging. We demonstrate that increasing levels of background fluorescence result in artifacts when using a linear perturbation algorithm, along with a significant loss of image fidelity and quantification accuracy. To correct for effects of background fluorescence, we have applied cubic polynomial fits to bulk raw measurements obtained from spatially homogeneous and heterogeneous phantoms. We show that subtraction of the average fluorescence response from the raw data before reconstruction can improve image quality and quantification accuracy as shown in relatively homogeneous or heterogeneous phantoms. Subtraction methods thus appear to be a promising route for adaptively correcting nonspecific background fluorochrome distribution.

Validation of in Vivo Fluorochrome Concentrations Measured Using Fluorescence Molecular Tomography

Journal of Biomedical Optics. Jul-Aug, 2005  |  Pubmed ID: 16178652

Fluorescence molecular tomography (FMT) has emerged as a means of quantitatively imaging fluorescent molecular probes in three dimensions in living systems. To assess the accuracy of FMT in vivo, translucent plastic tubes containing a turbid solution with a known concentration of Cy 5.5 fluorescent dye are constructed and implanted subcutaneously in nude mice, simulating the presence of a tumor accumulating a fluorescent molecular reporter. Comparisons between measurements of fluorescent tubes made before and after implantation demonstrate that the accuracy of FMT reported for homogeneous phantoms extends to the in vivo situation. The sensitivity of FMT to background fluorescence is tested by imaging fluorescent tubes in mice injected with Cy 5.5-labeled Annexin V. For small tube fluorochrome concentrations, the presence of background fluorescence results in increases in the reconstructed concentration. This phenomenon is counteracted by applying a simple subtraction correction to the measured fluorescence data. The effects of varying tumor photon absorption are simulated by imaging fluorescent tubes with varying ink concentrations, and are found to be minor. These findings demonstrate the in vivo quantitative accuracy of fluorescence tomography, and encourage further development of this imaging modality as well as application of FMT in molecular imaging studies using fluorescent reporters.

Use of Gene Expression Profiling to Direct in Vivo Molecular Imaging of Lung Cancer

Proceedings of the National Academy of Sciences of the United States of America. Oct, 2005  |  Pubmed ID: 16183744

Using gene expression profiling, we identified cathepsin cysteine proteases as highly up-regulated genes in a mouse model of human lung adenocarcinoma. Overexpression of cathepsin proteases in these lung tumors was confirmed by immunohistochemistry and Western blotting. Therefore, an optical probe activated by cathepsin proteases was selected to detect murine lung tumors in vivo as small as 1 mm in diameter and spatially separated. We generated 3D maps of the fluorescence signal and fused them with anatomical computed tomography images to show a close correlation between fluorescence signal and tumor burden. By serially imaging the same mouse, optical imaging was used to follow tumor progression. This study demonstrates the capability for molecular imaging of a primary lung tumor by using endogenous proteases expressed by a tumor. It also highlights the feasibility of using gene expression profiling to identify molecular targets for imaging lung cancer.

Accuracy of Fluorescent Tomography in the Presence of Heterogeneities: Study of the Normalized Born Ratio

IEEE Transactions on Medical Imaging. Oct, 2005  |  Pubmed ID: 16229423

We studied the performance of three-dimensional fluorescence tomography of diffuse media in the presence of heterogeneities. Experimental measurements were acquired using an imaging system consisting of a parallel plate-imaging chamber and a lens coupled charge coupled device camera, which enables conventional planar imaging as well as fluorescence tomography. To simulate increasing levels of background heterogeneity, we employed phantoms made of a fluorescent tube surrounded by several absorbers in different combinations of absorption distribution. We also investigated the effect of low absorbing thin layers (such as membranes). We show that the normalized Born approach accurately retrieves the position and shape of the fluorochrome even at high background heterogeneity. We also demonstrate that the quantification is relatively insensitive to a varying degree of heterogeneity and background optical properties. Findings are further contrasted to images obtained with the standard Born expansion and with a normalized approach that divides the fluorescent field with excitation measurements through a homogeneous medium.

Volumetric Tomography of Fluorescent Proteins Through Small Animals in Vivo

Proceedings of the National Academy of Sciences of the United States of America. Dec, 2005  |  Pubmed ID: 16344470

Volumetric detection and accurate quantification of fluorescent proteins in entire animals would greatly enhance our ability to monitor biological processes in vivo. Here we present a quantitative tomographic technique for visualization of superficial and deep-seated (>2-3 mm) fluorescent protein activity in vivo. We demonstrate noninvasive imaging of lung tumor progression in a murine model, as well as imaging of gene delivery using a herpes virus vector. This technology can significantly improve imaging capacity over the current state of the art and should find wide in vivo imaging applications in drug discovery, immunology, and cancer research.

Planar Fluorescence Imaging Using Normalized Data

Journal of Biomedical Optics. Nov-Dec, 2005  |  Pubmed ID: 16409072

Fluorescence imaging of tissues has gained significant attention in recent years due to the emergence of appropriate reporter technologies that enable noninvasive sensing of molecular function in vivo. Two major approaches have been used so far for fluorescence molecular imaging, i.e., epi-illumination (reflectance) imaging and fluorescence molecular tomography. Transillumination is an alternative approach that has been employed for imaging tissues in the past and could be similarly beneficial for fluorescence molecular imaging. We investigate data normalization schemes in reflectance and transillumination mode and experimentally demonstrate that normalized transillumination offers significant advantages over planar reflectance imaging and over nonnormalized methods. Our observations, based on phantoms and on postmortem and in vivo mouse measurements display image quality improvement, superior depth sensitivity, and improved imaging accuracy over the nonnormalized methods examined. Normalized planar imaging retains implementation simplicity and could be used to improve on standard fluorescence reflectance imaging and as a simplified alternative to the more integrated and accurate tomographic methods.

From Finite to Infinite Volumes: Removal of Boundaries in Diffuse Wave Imaging

Physical Review Letters. May, 2006  |  Pubmed ID: 16712298

In this Letter, we present a method that removes the contribution of the boundaries on the measurements from highly scattering media, transforming the signals captured from a bounded medium to measurements that would have been obtained if no boundary were present. This approach opens new possibilities in tomographic imaging in diffuse media as it eliminates the need for explicitly modeling boundaries and significantly simplifies reconstruction requirements.

Inflammation in Atherosclerosis: Visualizing Matrix Metalloproteinase Action in Macrophages in Vivo

Circulation. Jul, 2006  |  Pubmed ID: 16801460

Matrix metalloproteinases (MMPs) in inflamed atherosclerotic plaques may contribute to extracellular matrix remodeling and the onset of acute thrombotic complications.

Fluorescence Molecular Imaging

Annual Review of Biomedical Engineering. 2006  |  Pubmed ID: 16834550

There is a wealth of new fluorescent reporter technologies for tagging of many cellular and subcellular processes in vivo. This imposed contrast is now captured with an increasing number of available imaging methods that offer new ways to visualize and quantify fluorescent markers distributed in tissues. This is an evolving field of imaging sciences that has already achieved major advances but is also facing important challenges. It is nevertheless well poised to significantly impact the ways of biological research, drug discovery, and clinical practice in the years to come. Herein, the most pertinent technologies associated with in vivo noninvasive or minimally invasive fluorescence imaging of tissues are summarized. Focus is given to small-animal imaging. However, while a broad spectrum of fluorescence reporter technologies and imaging methods are outlined, as necessary for biomedical research, and clinical translation as well.

Fluorescence Molecular Tomography in the Presence of Background Fluorescence

Physics in Medicine and Biology. Aug, 2006  |  Pubmed ID: 16885619

Fluorescence molecular tomography is an emerging imaging technique that resolves the bio-distribution of engineered fluorescent probes developed for in vivo reporting of specific cellular and sub-cellular targets. The method can detect fluorochromes in picomole amounts or less, imaged through entire animals, but the detection sensitivity and imaging performance drop in the presence of background, non-specific fluorescence. In this study, we carried out a theoretical and an experimental investigation on the effect of background fluorescence on the measured signal and on the tomographic reconstruction. We further examined the performance of three subtraction methods based on physical models of photon propagation, using experimental data on phantoms and small animals. We show that the data pre-processing with subtraction schemes can improve image quality and quantification when non-specific background florescence is present.

Normalized Transillumination of Fluorescent Proteins in Small Animals

Molecular Imaging. Jul, 2006  |  Pubmed ID: 16954030

We investigated the capacity of normalized transillumination methods in imaging fluorescent proteins in visible light, in phantoms and in live mice. We demonstrate significant imaging improvements over epi-illumination imaging, as commonly applied today. Of particular importance is the significant betterment in contrast achieved, due to minimization of the influence of autofluorescence and the enhancement of depth sensitivity compared to epi-illumination imaging. Although normalized transillumination does not match the accuracy and quantification capacity of tomographic imaging, it nevertheless attains key advantages over other planar optical imaging methods while preserving implementation simplicity. Due to the balance achieved between simplicity and accuracy, normalized transillumination approaches could serve as an important alternative molecular imaging method.

Time-resolved Imaging of Optical Coefficients Through Murine Chest Cavities

Journal of Biomedical Optics. Nov-Dec, 2006  |  Pubmed ID: 17212540

As small animal optical imaging and tomography are gaining popularity for interrogating functional and molecular events in vivo, it becomes increasingly necessary to gain knowledge of the optical properties of the species investigated to better understand and describe photon propagation through their tissues. To achieve characterization of the spatial variation of average optical properties through murine chest cavities, time- and spatially resolved measurements of femto-second laser pulse transmission are performed through mice using a high-speed gated image intensifier. Application of time-resolved diffusion theory for finite slab geometry is first confirmed on phantoms and then applied to in vivo measurements for spatially resolving and quantifying mouse optical properties. Photon transmission images through mouse chest cavities are further obtained at different time gates to visualize the spatial variation observed and confirm the optical coefficient patterns calculated.

In-vivo Lung Cancer Imaging in Mice Using 360 Degrees Free-space Fluorescence Molecular Tomography

Conference Proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference. 2006  |  Pubmed ID: 17946109

We present the development and performance characteristics of a free-space fluorescence tomography system. The imaging system can capture complete angle projections of photons propagating through tissue in transillumination using a CCD camera. Experimental data on imaging lung cancer are presented. Overall, this imaging approach can offer unprecedented imaging performance in fluorescence molecular tomography of small animals.

Tomographic Fluorescence Imaging of Tumor Vascular Volume in Mice

Radiology. Mar, 2007  |  Pubmed ID: 17325064

To prospectively determine the feasibility of imaging vascular volume fraction (VVF) and its therapeutic inhibition in mouse models of cancer with three-dimensional fluorescence molecular tomography (FMT).

Fluorescence Tomography and Magnetic Resonance Imaging of Myocardial Macrophage Infiltration in Infarcted Myocardium in Vivo

Circulation. Mar, 2007  |  Pubmed ID: 17339546

Fluorescence imaging of the heart is currently limited to invasive ex vivo or in vitro applications. We hypothesized that the adaptation of advanced transillumination and tomographic techniques would allow noninvasive fluorescence images of the heart to be acquired in vivo and be coregistered with in vivo cardiac magnetic resonance images.

Free-space Fluorescence Molecular Tomography Utilizing 360 Degrees Geometry Projections

Optics Letters. Feb, 2007  |  Pubmed ID: 17356660

Fluorescence tomography of diffuse media can yield optimal three-dimensional imaging when multiple projections over 360 degrees geometries are captured, compared with limited projection angle systems such as implementations in the slab geometry. We demonstrate how it is possible to perform noncontact, 360 degrees projection fluorescence tomography of mice using CCD-camera-based detection in free space, i.e., in the absence of matching fluids. This approach achieves high spatial sampling of photons propagating through tissue and yields a superior information content data set compared with fiber-based 360 degrees implementations. Reconstruction feasibility using 36 projections in 10 degrees steps is demonstrated in mice.

Three-dimensional in Vivo Imaging of Green Fluorescent Protein-expressing T Cells in Mice with Noncontact Fluorescence Molecular Tomography

Molecular Imaging. Mar-Apr, 2007  |  Pubmed ID: 17445504

Given that optical tomography is capable of quantitatively imaging the distribution of several important chromophores and fluorophores in vivo, there has been a great deal of interest in developing optical imaging systems with increased numbers of measurements under optimal experimental conditions. In this article, we present a novel system that enables three-dimensional imaging of fluorescent probes in whole animals using a noncontact setup, in parallel with a three-dimensional surface reconstruction algorithm. This approach is directed toward the in vivo imaging of fluorophore or fluorescent protein concentration in small animals. The system consists of a rotating sample holder and a lens-coupled charge-coupled device camera in combination with a fiber-coupled laser scanning device. By measuring multiple projections, large data sets can be obtained, thus improving the accuracy of the inversion models used for quantitative three-dimensional reconstruction of fluorochrome distribution, as well as facilitating a higher spatial resolution. In this study, the system was applied to determining the distribution of green fluorescent protein (GFP)-expressing T lymphocytes in a transgenic mouse model, thus demonstrating the potential of the system for studying immune system function. The technique was used to image and reconstruct fluorescence originating from 32 x 10(6) T cells in the thymus and 3 x 10(5) T cells in the spleen.

Inversion with Early Photons

Medical Physics. Apr, 2007  |  Pubmed ID: 17500472

Optical tomography using early photons can improve resolution and reduce the ill-posed nature of the inversion problem. In this work we use 360 degrees projection experimental data to investigate the inversion performance of three commonly used numerical inversion methods: the random algebraic reconstruction technique (rART), singular value decomposition (SVD), and the conjugate-gradient-type method LSQR. Results are contrasted to each other and the effects of different photon propagation models are also investigated. We find that all methods perform adequately given appropriate regularization parameters, and that an experimentally measured photon weight function yields superior results over two approximate weights that have been previously used.

Noncontact Optical Imaging in Mice with Full Angular Coverage and Automatic Surface Extraction

Applied Optics. Jun, 2007  |  Pubmed ID: 17514324

During the past decade, optical imaging combined with tomographic approaches has proved its potential in offering quantitative three-dimensional spatial maps of chromophore or fluorophore concentration in vivo. Due to its direct application in biology and biomedicine, diffuse optical tomography (DOT) and its fluorescence counterpart, fluorescence molecular tomography (FMT), have benefited from an increase in devoted research and new experimental and theoretical developments, giving rise to a new imaging modality. The most recent advances in FMT and DOT are based on the capability of collecting large data sets by using CCDs as detectors, and on the ability to include multiple projections through recently developed noncontact approaches. For these to be implemented, we have developed an imaging setup that enables three-dimensional imaging of arbitrary shapes in fluorescence or absorption mode that is appropriate for small animal imaging. This is achieved by implementing a noncontact approach both for sources and detectors and coregistering surface geometry measurements using the same CCD camera. A thresholded shadowgrammetry approach is applied to the geometry measurements to retrieve the surface mesh. We present the evaluation of the system and method in recovering three-dimensional surfaces from phantom data and live mice. The approach is used to map the measured in vivo fluorescence data onto the tissue surface by making use of the free-space propagation equations, as well as to reconstruct fluorescence concentrations inside highly scattering tissuelike phantom samples. Finally, the potential use of this setup for in vivo small animal imaging and its impact on biomedical research is discussed.

Optimization of 360 Degrees Projection Fluorescence Molecular Tomography

Medical Image Analysis. Aug, 2007  |  Pubmed ID: 17524701

Fluorescence tomography of tissues has been generally limited to systems that require fixed geometries or measurements employing fibers. Certain technological advances however, have more recently allowed the development of complete-projection 360 degrees tomographic approaches using non-contact detection and illumination. Employing multiple illumination projections and CCD cameras as detection devices vastly increases the information content acquired, posing non-trivial computational and experimental requirements. In this paper, we use singular-value analysis to optimize experimental parameters relevant to the design and operation of emerging 360 degrees fluorescence molecular tomography (FMT) methods and systems for small animal imaging. We present the theoretical and experimental methodology, optimization results and their experimental validation. We further discuss how these results can be employed to improve the performance of existing FMT systems and guide the design of new systems.

A Statistical Approach to Inverting the Born Ratio

IEEE Transactions on Medical Imaging. Jul, 2007  |  Pubmed ID: 17649903

We examine the problem of fluorescence molecular tomography using the normalized Born approximation, termed herein the Born ratio, from a statistical perspective. Experimentally verified noise models for received signals at the excitation and emission wavelengths are combined to generate a stochastic model for the Born ratio. This model is then utilized within a maximum likelihood framework to obtain an inverse solution based on a fixed point iteration. Results are presented for three experimental scenarios: phantom data with a homogeneous background, phantoms implanted within a small animal, and in vivo data using an exogenous probe.

Multispectral Photoacoustic Imaging of Fluorochromes in Small Animals

Optics Letters. Oct, 2007  |  Pubmed ID: 17909608

Fluorochromes have become essential reporter molecules in biological research. We show that the depth-resolved distribution of fluorochromes in small animals can be imaged with 25 fmol sensitivity and 150 microm spatial resolution by means of multispectral photoacoustic imaging. The major advantage of the multispectral approach is the sensitive differentiation of chromophores and fluorochromes of interest based on self-reference measurements, as evidenced in this study by resolving a commonly used fluorochrome (Alexa Fluor 750) in mouse. The suggested method is well suited for enhancing visualization of functional and molecular information in vivo and longitudinally.

Hybrid Photoacoustic Fluorescence Molecular Tomography Using Finite-element-based Inversion

Medical Physics. Nov, 2007  |  Pubmed ID: 18072494

Improvements in fluorescence reconstruction when utilizing a hybrid photoacoustic (PAT) fluorescence molecular tomography (FMT) method to image optically heterogeneous media are studied and showcased. Quantitative optical absorption maps are retrieved using a normalized backprojection algorithm for PAT reconstruction. Consecutively, the reconstructed absorption distribution is employed into computing a diffusion-equation-based forward model for FMT using a finite-element solution. The potential promise of the suggested method is experimentally verified on tissue-mimicking fluorescent phantoms, where improvements in the quality of FMT reconstructions are observed when imaging at the presence of a large absorber.

Development of Fluorescent Materials for Diffuse Fluorescence Tomography Standards and Phantoms

Optics Express. Jul, 2007  |  Pubmed ID: 19547203

The availability of fluorescence standards is necessary in the development of systems and methods for fluorescence imaging. In this study, two approaches for developing diffuse fluorescence materials to be used as standards or phantoms in diffuse fluorescent tomography applications were investigated. Specifically, silicone rubber and polyester casting resin were used as base materials, and silicone pigments or TiO(2) / India Ink were added respectively to vary the optical properties. Characterization of the optical properties achieved was performed using time-resolved methods. Subsequently, different near-infrared fluorochromes were examined for imparting controlled and stable fluorescence properties. It was determined that hydrophobic fluorophores (IR 676 and IR 780 Iodide) suspended in dichloromethane and hydrophilic fluorophores (Cy5.5 and AF 750) suspended in methanol produced diffusive silicone and resin fluorescent materials, respectively. However only the hydrophobic fluorophores embedded within silicone resulted in the construction of a material with the characteristics of a standard, i.e. stability of fluorescence intensity with time and a linear dependence of normalized fluorescence intensity to fluorophore concentration.

In Vivo Imaging of Drosophila Melanogaster Pupae with Mesoscopic Fluorescence Tomography

Nature Methods. Jan, 2008  |  Pubmed ID: 18066071

We report a technique for fluorescence tomography that operates beyond the penetration limits of tissue-sectioning fluorescence microscopy. The method uses multi-projection illumination and photon transport description in opaque tissues. We demonstrate whole-body three-dimensional visualization of the morphogenesis of GFP-expressing salivary glands and wing imaginal discs in living Drosophila melanogaster pupae in vivo and over time.

3D Multi-modal Registration for Assessing Molecular Activity Changes in Time-dependent Geometries

Conference Proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference. 2008  |  Pubmed ID: 19163583

An important issue for in-vivo fluorescence molecular imaging and tomography has been the robust registration of animals imaged at different time points in order to improve the accuracy of monitoring fluorophore distribution. This paper presents a novel automatic method for aligning 3D temporal data of small animals based on the robust detection of surface anatomical features. Since it is based on 3D surface landmarks the method can be used for mono-modal or multi-modal X-ray CT and/or Optical tomography data in order to account for differences in the positioning and compression of small animals. It can be further extended to other imaging modalities, as well.

Visualization of Pulmonary Inflammation Using Noninvasive Fluorescence Molecular Imaging

Journal of Applied Physiology (Bethesda, Md. : 1985). Mar, 2008  |  Pubmed ID: 18202169

The ability to visualize molecular processes and cellular regulators of complex pulmonary diseases such as asthma, chronic obstructive pulmonary disease (COPD), or adult respiratory distress syndrome (ARDS), would aid in the diagnosis, differentiation, therapy assessment and in small animal-based drug-discovery processes. Herein we report the application of normalized transillumination and fluorescence molecular tomography (FMT) for the noninvasive quantitative imaging of the mouse lung in vivo. We demonstrate the ability to visualize and quantitate pulmonary response in a murine model of LPS-induced airway inflammation. Twenty-four hours prior to imaging, BALB/c female mice were injected via tail vein with 2 nmol of a cathepsin-sensitive activatable fluorescent probe (excitation: 750 nm; emission: 780 nm) and 2 nmol of accompanying intravascular agent (excitation: 674 nm; emission: 694 nm). Six hours later, the mice were anesthetized with isoflurane and administered intranasal LPS in sterile 0.9% saline in 25 microl aliquots (one per nostril). Fluorescence molecular imaging revealed the in vivo profile of cysteine protease activation and vascular distribution within the lung typifying the inflammatory response to LPS insult. Results were correlated with standard in vitro laboratory tests (Western blot, bronchoalveolar lavage or BAL analysis, immunohistochemistry) and revealed good correlation with the underlying activity. We demonstrated the capacity of fluorescence tomography to noninvasively and longitudinally characterize physiological, cellular, and subcellular processes associated with inflammatory disease burden in the lung. The data presented herein serve to further evince fluorescence molecular imaging as a technology highly appropriate for the biomedical laboratory.

Surface Reconstruction for Free-space 360 Degrees Fluorescence Molecular Tomography and the Effects of Animal Motion

IEEE Transactions on Medical Imaging. Feb, 2008  |  Pubmed ID: 18334440

Complete projection (360 degrees ) free-space fluorescence tomography of opaque media is poised to enable 3-D imaging through entire small animals in vivo with improved depth resolution compared to 360 degrees -projection fiber-based systems or limited-view angle systems. This approach can lead to a new generation of Fluorescence Molecular Tomography (FMT) performance since it allows high spatial sampling of photon fields propagating through tissue at any projection, employing nonconstricted animal surfaces. Herein, we employ a volume carving method to capture 3-D surfaces of diffusive objects and register the captured surface in the geometry of an FMT 360 degrees -projection acquisition system to obtain 3-D fluorescence image reconstructions. Using experimental measurements we evaluate the accuracy of the surface capture procedure by reconstructing the surfaces of phantoms of known dimensions. We then employ this methodology to characterize the animal movement of anaesthetized animals. We find that the effects of animal movement on the FMT reconstructed image were within system resolution limits (approximately 0.07 cm).

Multispectral Imaging Using Multiple-bandpass Filters

Optics Letters. May, 2008  |  Pubmed ID: 18451974

We present a method for multispectral imaging. This method uses color CCD cameras with a multiple-bandpass filter, which modifies the spectral response of the cameras used and enables concurrent acquisition of multiple images at defined spectral bands. We experimentally demonstrate methodological feasibility using two color CCD cameras and a polychroic mirror to simultaneously capture eight spectral bands. We discuss how the method developed is well suited for multispectral applications of transient phenomena or for real-time measurements.

Small-animal Optical Imaging

Journal of Biomedical Optics. Jan-Feb, 2008  |  Pubmed ID: 18456953

Investigating Pharmacology in Vivo Using Magnetic Resonance and Optical Imaging

Drugs in R&D. 2008  |  Pubmed ID: 18720999

The better and earlier a disease can be diagnosed and characterized, the greater the chance of being able to intervene in this process with a chemical entity. This is the rationale for the use of in vivo imaging techniques in the drug discovery and development process. In this article we address the value of two imaging modalities in this area, i.e. magnetic resonance imaging (MRI) and optical imaging. The multiparametric nature of MRI enables anatomical, functional, metabolic and, to a certain extent, also cellular and target-related information to be obtained noninvasively at high spatial resolution. This favours characterization of a disease state and the corresponding drug intervention. The noninvasiveness of MRI strengthens the link between preclinical and clinical pharmaceutical research. The high sensitivity of optical techniques enables molecular information to be obtained in vivo. Within pharmacological research, the main applications of optical techniques relate to the early drug discovery process and acquisition of target-related information. However, potential clinical applications of optical imaging are also emerging. The complementary character of both imaging modalities renders them useful in various portions of the drug discovery process, from early target selection and validation to clinical studies.

Registration of 3-D CT and 2-D Flat Images of Mouse Via Affine Transformation

IEEE Transactions on Information Technology in Biomedicine : a Publication of the IEEE Engineering in Medicine and Biology Society. Sep, 2008  |  Pubmed ID: 18779071

It is difficult to directly coregister the 3-D fluorescence molecular tomography (FMT) image of a small tumor in a mouse whose maximal diameter is only a few millimeters with a larger CT image of the entire animal that spans about 10 cm. This paper proposes a new method to register 2-D flat and 3-D CT image first to facilitate the registration between small 3-D FMT images and large 3-D CT images. A novel algorithm combining differential evolution and improved simplex method for the registration between the 2-D flat and 3-D CT images is introduced and validated with simulated images and real images of mice. The visualization of the alignment of the 3-D FMT and CT image through 2-D registration shows promising results.

Real-time Catheter Molecular Sensing of Inflammation in Proteolytically Active Atherosclerosis

Circulation. Oct, 2008  |  Pubmed ID: 18852366

To enable intravascular detection of inflammation in atherosclerosis, we developed a near-infrared fluorescence (NIRF) catheter-based strategy to sense cysteine protease activity during vascular catheterization.

Polarization-sensitive Optoacoustic Tomography of Optically Diffuse Tissues

Optics Letters. Oct, 2008  |  Pubmed ID: 18923605

Polarization is indicative of material anisotropy, a property that reveals structural orientation information of molecules inside the material. Herein we investigate whether polarization can be detected optoacoustically in scattering and absorbing tissues. Using a laboratory prototype of polarization-sensitive optoacoustic tomography, we demonstrate high-resolution reconstructions of dichroism contrast deep in optically diffusive tissue-mimicking phantoms. The technique is expected to enable highly accurate imaging of polarization contrast in tissues, far beyond the current capabilities of pure optical polarization-imaging approaches.

Early Photon Tomography Allows Fluorescence Detection of Lung Carcinomas and Disease Progression in Mice in Vivo

Proceedings of the National Academy of Sciences of the United States of America. Dec, 2008  |  Pubmed ID: 19015534

Imaging of targeted fluorescent probes offers significant advantages for investigating disease and tissue function in animal models in vivo. Conversely, macroscopic tomographic imaging is challenging because of the high scatter of light in biological tissue and the ill-posed nature of the reconstruction mathematics. In this work, we use the earliest-transmitted photons through Lewis Lung Carcinoma bearing mice, thereby dramatically reducing the effect of tissue scattering. By using a fluorescent probe sensitive to cysteine proteases, the method yielded outstanding imaging performance compared with conventional approaches. Accurate visualization of biochemical abnormalities was achieved, not only in the primary tumor, but also in the surrounding tissue related to cancer progression and inflammatory response at the organ level. These findings were confirmed histologically and with ex vivo fluorescence microscopy. The imaging fidelity demonstrated underscores a method that can use a wide range of fluorescent probes to accurately visualize cellular- and molecular-level events in whole animals in vivo.

Performance of the Red-shifted Fluorescent Proteins in Deep-tissue Molecular Imaging Applications

Journal of Biomedical Optics. Jul-Aug, 2008  |  Pubmed ID: 19021336

The discovery of new fluorescent proteins (FPs) that emit in the far-red part of the spectrum, where light absorption from tissue is significantly lower than in the visible, offers the possibility for noninvasive biological interrogation at the entire organ or small animal level in vivo. The performance of FPs in deep-tissue imaging depends not only on their optical characteristics, but also on the wavelength-dependent tissue absorption and the depth of the fluorescence activity. To determine the optimal choice of FP and illumination wavelength, we compared the performance of five of the most promising FPs: tdTomato, mCherry, mRaspberry, mPlum, and Katushka. We experimentally measured the signal strength through mice and employed theoretical predictions to obtain an understanding of the performance of different illumination scenarios, especially as they pertain to tomographic imaging. It was found that the appropriate combination of red-shifted proteins and illumination wavelengths can improve detection sensitivity in small animals by at least two orders of magnitude compared with green FP. It is also shown that the steep attenuation change of the hemoglobin spectrum around the 600-nm range may significantly affect the detection sensitivity and, therefore, necessitates the careful selection of illumination wavelengths for optimal imaging performance.

Hybrid FMT-CT Imaging of Amyloid-beta Plaques in a Murine Alzheimer's Disease Model

NeuroImage. Feb, 2009  |  Pubmed ID: 19041402

The need to study molecular and functional parameters of Alzheimer's disease progression in animal models has led to the development of disease-specific fluorescent markers. However, curved optical interfaces and a highly heterogeneous internal structure make quantitative fluorescence imaging of the murine brain a particularly challenging tomographic problem. We investigated the integration of X-ray computed tomography (CT) information into a state-of-the-art fluorescence molecular tomography (FMT) scheme and establish that the dual-modality approach is essential for high fidelity reconstructions of distributed fluorescence within the murine brain, as compared to conventional fluorescence tomography. We employ this method in vivo using a fluorescent oxazine dye to quantify amyloid-beta plaque burden in transgenic APP23 mice modeling Alzheimer's disease. Multi-modal imaging allows for accurate signal localization and correlation of in vivo findings to ex vivo studies. The results point to FMT-CT as an essential tool for in vivo study of neurodegenerative disease in animal models and potentially humans.

Normalized Born Ratio for Fluorescence Optical Projection Tomography

Optics Letters. Feb, 2009  |  Pubmed ID: 19183644

We present a normalized Born approach for fluorescence optical projection tomography that takes into account tissue absorption properties. This approach can be particularly useful to study fluorochrome distribution within tissue. We use the algorithm to three-dimensionally reconstruct and characterize a fluorescein isothiocyanate containing absorptive phantom and an infarcted mouse heart previously injected with a fluorescent molecular probe.

In Vivo Investigation of Breast Cancer Progression by Use of an Internal Control

Neoplasia (New York, N.Y.). Mar, 2009  |  Pubmed ID: 19242603

Optical imaging of breast cancer has been considered for detecting functional and molecular characteristics of diseases in clinical and preclinical settings. Applied to laboratory research, photonic investigations offer a highly versatile tool for preclinical imaging and drug discovery. A particular advantage of the optical method is the availability of multiple spectral bands for performing imaging. Herein, we capitalize on this feature to demonstrate how it is possible to use different wavelengths to offer internal controls and significantly improve the observation accuracy in molecular imaging applications. In particular, we show the independent in vivo detection of cysteine proteases along with tumor permeability and interstitial volume measurements using a dual-wavelength approach. To generate results with a view toward clinically geared studies, a transgenic Her2/neu mouse model that spontaneously developed mammary tumors was used. In vivo findings were validated against conventional ex vivo tests such as histology and Western blot analyses. By correcting for biodistribution parameters, the dual-wavelength method increases the accuracy of molecular observations by separating true molecular target from probe biodistribution. As such, the method is highly appropriate for molecular imaging studies where often probe delivery and target presence are not independently assessed. On the basis of these findings, we propose the dual-wavelength/normalization approach as an essential method for drug discovery and preclinical imaging studies.

Performance Dependence of Hybrid X-ray Computed Tomography/fluorescence Molecular Tomography on the Optical Forward Problem

Journal of the Optical Society of America. A, Optics, Image Science, and Vision. Apr, 2009  |  Pubmed ID: 19340266

Hybrid imaging systems combining x-ray computed tomography (CT) and fluorescence tomography can improve fluorescence imaging performance by incorporating anatomical x-ray CT information into the optical inversion problem. While the use of image priors has been investigated in the past, little is known about the optimal use of forward photon propagation models in hybrid optical systems. In this paper, we explore the impact on reconstruction accuracy of the use of propagation models of varying complexity, specifically in the context of these hybrid imaging systems where significant structural information is known a priori. Our results demonstrate that the use of generically known parameters provides near optimal performance, even when parameter mismatch remains.

Imaging of Mesoscopic-scale Organisms Using Selective-plane Optoacoustic Tomography

Physics in Medicine and Biology. May, 2009  |  Pubmed ID: 19369709

Mesoscopic-scale living organisms (i.e. 1 mm to 1 cm sized) remain largely inaccessible by current optical imaging methods due to intensive light scattering in tissues. Therefore, imaging of many important model organisms, such as insects, fishes, worms and similarly sized biological specimens, is currently limited to embryonic or other transparent stages of development. This makes it difficult to relate embryonic cellular and molecular mechanisms to consequences in organ function and animal behavior in more advanced stages and adults. Herein, we have developed a selective-plane illumination optoacoustic tomography technique for in vivo imaging of optically diffusive organisms and tissues. The method is capable of whole-body imaging at depths from the sub-millimeter up to centimeter range with a scalable spatial resolution in the order of magnitude of a few tenths of microns. In contrast to pure optical methods, the spatial resolution here is not determined nor limited by light diffusion; therefore, such performance cannot be achieved by any other optical imaging technology developed so far. The utility of the method is demonstrated on several whole-body models and small-animal extremities.

Sensitivity of Molecular Target Detection by Multispectral Optoacoustic Tomography (MSOT)

Medical Physics. Mar, 2009  |  Pubmed ID: 19378754

Optoacoustic imaging is emerging as a noninvasive imaging modality that can resolve optical contrast through several millimeters to centimeters of tissue with the resolution achieved by ultrasound imaging. More recently, applied at multiple illumination wavelengths, multispectral optoacoustic tomography (MSOT) offered the ability to effectively visualize tissue biomarkers by resolving their distinct spectral signatures. While the imaging potential of the method has been demonstrated, little is known on the sensitivity performance in resolving chromophoric and fluorescent substances, such as optical functional and molecular reporters. Herein the authors investigate the detection capacity and physical limits of tomographic optoacoustic imaging by simulating signals originating from absorbing spheres in tissue-mimicking media. To achieve this, a modified optoacoustic equation is employed to incorporate wavelength-dependent propagation and attenuation of diffuse light and ultrasound. The theoretical predictions are further validated in phantom experiments involving Cy5.5, a common near-infrared fluorescent molecular agent.

Surface Modification and Size Dependence in Particle Translocation During Early Embryonic Development

Inhalation Toxicology. Jul, 2009  |  Pubmed ID: 19558239

Since the mid-1990 s, the number of studies linking air pollutants to preterm and low birth weight, as well as to cardiac birth defects, has grown steadily each year. The critical period in the development of mouse embryos begins with the commencement of gastrulation at day 7.5 of gestation. Our aim is to examine the role of particles size and surface modification in particle translocation during this early embryonic development. Fluorescent polystyrene particles (PS) were employed because they offer an efficient and safe tracking method. Pregnant female mice were sacrificed at 7.5 days of gestation. After cutting open the deciduas, the parietal endoderm was carefully separated and removed. Different sizes of amine- and carboxyl-modified PS beads were injected via the extraembryonic tissue. The embryos were incubated for 12 h, and were investigated under fluorescent microscopy, confocal microscopy, and mesoscopic fluorescence tomography. The results show that 20-nm carboxylic PS distribute in the embryonic and extraembryonic germ layers of ectoderm, mesoderm, and endoderm. Moreover, when the particles are bigger than 100 nm, PS accumulate in extraembryonic tissue, but nevertheless 200-nm amine-modified particles can pass into the embryos. Interestingly, a growth inhibition was observed in the embryos containing nanoparticles. Finally, the stronger translocation effect is associated with amine-modified PS beads (200 nm) instead of the smaller (20 nm, 100 nm) carboxyl ones.

In-vivo Imaging of Murine Tumors Using Complete-angle Projection Fluorescence Molecular Tomography

Journal of Biomedical Optics. May-Jun, 2009  |  Pubmed ID: 19566290

We interrogate the ability of free-space fluorescence tomography to image small animals in vivo using charge-coupled device (CCD) camera measurements over 360-deg noncontact projections. We demonstrate the performance of normalized dual-wavelength measurements that are essential for in-vivo use, as they account for the heterogeneous distribution of photons in tissue. In-vivo imaging is then showcased on mouse lung and brain tumors cross-validated by x-ray microcomputed tomography and histology.

Quantitative Optoacoustic Signal Extraction Using Sparse Signal Representation

IEEE Transactions on Medical Imaging. Dec, 2009  |  Pubmed ID: 19628454

We report on a new quantification methodology of optoacoustic tomographic reconstructions under heterogeneous illumination conditions representative of realistic whole-body imaging scenarios. Our method relies on the differences in the spatial characteristics of the absorption coefficient and the optical energy density within the medium. By using sparse-representation based decomposition, we exploit these different characteristics to extract both the absorption coefficient and the photon density within the imaged object from the optoacoustic image. In contrast to previous methods, this algorithm is not based on the solution of theoretical light transport equations and it does not require explicit knowledge of the illumination geometry or the optical properties of the object and other unknown or loosely defined experimental parameters, leading to highly robust performance. The method was successfully examined with numerically and experimentally generated data and was found to be ideally suited for practical implementations in tomographic schemes of varying complexity, including multiprojection illumination systems and multispectral optoacoustic tomography (MSOT) studies of tissue biomarkers.

Optical Imaging of Molecular Signatures in Pulmonary Inflammation

Proceedings of the American Thoracic Society. Aug, 2009  |  Pubmed ID: 19687213

Biomedical imaging has become an important tool in the study of "-omics" fields by allowing the noninvasive visualization of functional and molecular events using in vivo staining and reporter gene approaches. This capacity can go beyond the understanding of the genetic basis and phenotype of such respiratory conditions as acute bronchitis, adult respiratory distress syndrome (ARDS), chronic obstructive pulmonary disease (COPD), and asthma and investigate the development of disease and of therapeutic events longitudinally and in unperturbed environments. Herein, we show how the application of novel quantitative optical imaging methods, using transillumination and fluorescence molecular tomography (FMT), can allow visualization of pulmonary inflammation in small animals in vivo. The results confirm prior observations using a protease-sensitive probe. We discuss how this approach enables in vivo insights at the system level as to the dynamic role of proteases in respiratory pathophysiology and their potential as therapeutic targets. Overall, the proposed imaging method can be used with a significantly wider range of possible targets and applications in lung imaging.

Transillumination Fluorescence Imaging in Mice Using Biocompatible Upconverting Nanoparticles

Optics Letters. Sep, 2009  |  Pubmed ID: 19724491

We report on a systematic study of upconverting fluorescence signal generation within turbid phantoms and real tissues. An accurate three-point Green's function, describing the forward model of photon propagation, is established and experimentally validated. We further demonstrate, for the first time to our knowledge, autofluorescence-free transillumination imaging of mice that have received biocompatible upconverting nanoparticles. The method holds great promise for artifact-free whole-body visualization of optical molecular probes.

Complete Angle Small Animal Fluorescence Imaging with Early-arriving Photons

Conference Proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference. 2009  |  Pubmed ID: 19964155

Fluorescence mediated tomography allows quantitative, three-dimensional imaging of optical reporter probes in whole animals and is therefore emerging as a powerful molecular imaging tool. The achievable image quality in fluorescence tomography is limited by the high-degree of light scatter in biological tissue. Time-gated detection of early-arriving and therefore minimally-scattered photons transmitted through diffusive tissue is one strategy for minimizing the effects of light scatter. In this work, we performed full-angle tomographic imaging of mice implanted with fluorescent tubes using time-gated detection of early- and later-arriving photons. This was achieved using a femtosecond laser and a high-speed, time-gated intensified CCD imager. We demonstrate that the early-transmitted fluorescent photons allow improved visualization of the fluorescence distribution, even when considering individual projections through the animal. High-fidelity image reconstruction using 72 projections in 5-degree steps using early-arriving photons is also demonstrated.

Multispectral Optoacoustic Tomography (MSOT) Scanner for Whole-body Small Animal Imaging

Optics Express. Nov, 2009  |  Pubmed ID: 19997381

A major difficulty arising from whole-body optoacoustic imaging is the long acquisition times associated with recording signals from multiple spatial projections. The acquired signals are also generally weak and the signal-to-noise-ratio is low, problems often solved by signal averaging, which complicates acquisition and increases acquisition times to an extent that makes many in vivo applications challenging or even impossible. Herein we present a fast acquisition multispectral optoacoustic tomography (MSOT) scanner for whole-body visualization of molecular markers in small animals. Multi-wavelength illumination offers the possibility to resolve exogenously administered fluorescent probes, biomarkers, and other intrinsic and exogenous chromophores. The system performance is determined in phantom experiments involving molecular probes and validated by imaging of small animals of various scales.

Real-time Intraoperative Fluorescence Imaging System Using Light-absorption Correction

Journal of Biomedical Optics. Nov-Dec, 2009  |  Pubmed ID: 20059250

We present a novel fluorescence imaging system developed for real-time interventional imaging applications. The system implements a correction scheme that improves the accuracy of epi-illumination fluorescence images for light intensity variation in tissues. The implementation is based on the use of three cameras operating in parallel, utilizing a common lens, which allows for the concurrent collection of color, fluorescence, and light attenuation images at the excitation wavelength from the same field of view. The correction is based on a ratio approach of fluorescence over light attenuation images. Color images and video is used for surgical guidance and for registration with the corrected fluorescence images. We showcase the performance metrics of this system on phantoms and animals, and discuss the advantages over conventional epi-illumination systems developed for real-time applications and the limits of validity of corrected epi-illumination fluorescence imaging.

Comparison of Fluorescence Tomographic Imaging in Mice with Early-arriving and Quasi-continuous-wave Photons

Optics Letters. Feb, 2010  |  Pubmed ID: 20125724

The highly diffuse nature of light propagation in biological tissue is a major challenge for obtaining high-fidelity fluorescence tomographic images. In this work we investigated the use of time-gated detection of early-arriving photons for reducing the effects of light scatter in mice relative to quasi-cw photons. When analyzing sinographic representations of the measured data, it was determined that early photons allowed a reduction in the measured FWHM of fluorescent targets by a factor of approximately 2-3, yielding a significant improvement in the tomographic image reconstruction quality.

Multifunctional Nanocarriers for Diagnostics, Drug Delivery and Targeted Treatment Across Blood-brain Barrier: Perspectives on Tracking and Neuroimaging

Particle and Fibre Toxicology. 2010  |  Pubmed ID: 20199661

Nanotechnology has brought a variety of new possibilities into biological discovery and clinical practice. In particular, nano-scaled carriers have revolutionalized drug delivery, allowing for therapeutic agents to be selectively targeted on an organ, tissue and cell specific level, also minimizing exposure of healthy tissue to drugs. In this review we discuss and analyze three issues, which are considered to be at the core of nano-scaled drug delivery systems, namely functionalization of nanocarriers, delivery to target organs and in vivo imaging. The latest developments on highly specific conjugation strategies that are used to attach biomolecules to the surface of nanoparticles (NP) are first reviewed. Besides drug carrying capabilities, the functionalization of nanocarriers also facilitate their transport to primary target organs. We highlight the leading advantage of nanocarriers, i.e. their ability to cross the blood-brain barrier (BBB), a tightly packed layer of endothelial cells surrounding the brain that prevents high-molecular weight molecules from entering the brain. The BBB has several transport molecules such as growth factors, insulin and transferrin that can potentially increase the efficiency and kinetics of brain-targeting nanocarriers. Potential treatments for common neurological disorders, such as stroke, tumours and Alzheimer's, are therefore a much sought-after application of nanomedicine. Likewise any other drug delivery system, a number of parameters need to be registered once functionalized NPs are administered, for instance their efficiency in organ-selective targeting, bioaccumulation and excretion. Finally, direct in vivo imaging of nanomaterials is an exciting recent field that can provide real-time tracking of those nanocarriers. We review a range of systems suitable for in vivo imaging and monitoring of drug delivery, with an emphasis on most recently introduced molecular imaging modalities based on optical and hybrid contrast, such as fluorescent protein tomography and multispectral optoacoustic tomography. Overall, great potential is foreseen for nanocarriers in medical diagnostics, therapeutics and molecular targeting. A proposed roadmap for ongoing and future research directions is therefore discussed in detail with emphasis on the development of novel approaches for functionalization, targeting and imaging of nano-based drug delivery systems, a cutting-edge technology poised to change the ways medicine is administered.

Intravascular Near-infrared Fluorescence Molecular Imaging of Atherosclerosis: Toward Coronary Arterial Visualization of Biologically High-risk Plaques

Journal of Biomedical Optics. Jan-Feb, 2010  |  Pubmed ID: 20210433

New imaging methods are urgently needed to identify high-risk atherosclerotic lesions prior to the onset of myocardial infarction, stroke, and ischemic limbs. Molecular imaging offers a new approach to visualize key biological features that characterize high-risk plaques associated with cardiovascular events. While substantial progress has been realized in clinical molecular imaging of plaques in larger arterial vessels (carotid, aorta, iliac), there remains a compelling, unmet need to develop molecular imaging strategies targeted to high-risk plaques in human coronary arteries. We present recent developments in intravascular near-IR fluorescence catheter-based strategies for in vivo detection of plaque inflammation in coronary-sized arteries. In particular, the biological, light transmission, imaging agent, and engineering principles that underlie a new intravascular near-IR fluorescence sensing method are discussed. Intravascular near-IR fluorescence catheters appear highly translatable to the cardiac catheterization laboratory, and thus may offer a new in vivo method to detect high-risk coronary plaques and to assess novel atherosclerosis biologics.

Fast Semi-analytical Model-based Acoustic Inversion for Quantitative Optoacoustic Tomography

IEEE Transactions on Medical Imaging. Jun, 2010  |  Pubmed ID: 20304725

We present a fast model-based inversion algorithm for quantitative 2-D and 3-D optoacoustic tomography. The algorithm is based on an accurate and efficient forward model, which eliminates the need for regularization in the inversion process while providing modeling flexibility essential for quantitative image formation. The resulting image-reconstruction method eliminates stability problems encountered in previously published model-based techniques and, thus, enables performing image reconstruction in real time. Our model-based framework offers a generalization of the forward solution to more comprehensive optoacoustic propagation models, such as including detector frequency response, without changing the inversion procedure. The reconstruction speed and other algorithmic performances are demonstrated using numerical simulation studies and experimentally on tissue-mimicking optically heterogeneous phantoms and small animals. In the experimental examples, the model-based reconstructions manifested correctly the effect of light attenuation through the objects and did not suffer from the artifacts which usually afflict the commonly used filtered backprojection algorithms, such as negative absorption values.

Imaging of Molecular Probe Activity with Born-normalized Fluorescence Optical Projection Tomography

Optics Letters. Apr, 2010  |  Pubmed ID: 20364226

Optical projection tomography is a new ex vivo imaging technique that allows imaging of whole organs in three dimensions at high spatial resolutions. In this Letter we demonstrate its capability to tomographically visualize molecular activity in whole organs of mice. In particular, eosinophil activity in asthmatic lungs is resolved using a Born-normalized fluorescence optical projection tomography and employing a near-IR molecular probe. The possibility to achieve molecularly sensitive imaging contrast in optical projection tomography by means of targeted and activatable imaging reporter agents adds a new range of capabilities for investigating molecular signatures of pathophysiological processes and a wide variety of diseases and their development.

Molecular Imaging by Means of Multispectral Optoacoustic Tomography (MSOT)

Chemical Reviews. May, 2010  |  Pubmed ID: 20387910

Imaging Performance of a Hybrid X-ray Computed Tomography-fluorescence Molecular Tomography System Using Priors

Medical Physics. May, 2010  |  Pubmed ID: 20527531

The performance is studied of two newly introduced and previously suggested methods that incorporate priors into inversion schemes associated with data from a recently developed hybrid x-ray computed tomography and fluorescence molecular tomography system, the latter based on CCD camera photon detection. The unique data set studied attains accurately registered data of high spatially sampled photon fields propagating through tissue along 360 degrees projections.

Mesoscopic Epifluorescence Tomography: Reconstruction of Superficial and Deep Fluorescence in Highly-scattering Media

Optics Express. Apr, 2010  |  Pubmed ID: 20588688

Mesoscopic Epifluorescence Tomography (MEFT) is a technique derived from Laminar Optical Tomography (LOT), determining fluorescence biodistribution by tomographic means in reflectance geometry. A pencil beam is scanned over the region of interest to excite fluorophores hidden within the tissue, while a CCD camera acquires images of reflected fluorescence emissions. This configuration is advantageous whenever transillumination of the specimen is not feasible, e.g., in the presence of skin chambers or when using wavelengths in the visible range where absorption is high. We present simulation and phantom studies recovering deep GFP-like fluorescence in highly scattering and strongly absorbing media with a penetration depth up to 10mm.

Near-infrared Fluorescence Catheter System for Two-dimensional Intravascular Imaging in Vivo

Optics Express. May, 2010  |  Pubmed ID: 20588998

Detection of high-risk coronary arterial plaques prior to rupture remains an unmet clinical challenge, in part due to the stringent resolution and sensitivity requirements for in vivo human coronary arterial imaging. To address this need, we have developed a near-infrared (NIR) fluorescence imaging catheter system for intra-vascular molecular imaging of atherosclerosis in coronary artery-sized vessels, capable of resolving two-dimensional fluorescence activity in hollow organs, such as blood vessels. Based on a rotational fiber design, the catheter system illuminates and detects perpendicular to the rotational axis, while an automated pullback mechanism enables visualization along blood vessels with a scan speed of up to 1.5 mm/sec. We demonstrate the previously undocumented capacity to produce intravascular NIR fluorescence images of hollow organs in vivo and showcase the performance metrics of the system developed using blood vessel mimicking phantoms. This imaging approach is geared toward in vivo molecular imaging of atherosclerotic biomarkers and is engineered to allow seamless integration into the cardiac catheterization laboratory.

Fast Automatic Segmentation of Anatomical Structures in X-ray Computed Tomography Images to Improve Fluorescence Molecular Tomography Reconstruction

Journal of Biomedical Optics. May-Jun, 2010  |  Pubmed ID: 20615008

The recent development of hybrid imaging scanners that integrate fluorescence molecular tomography (FMT) and x-ray computed tomography (XCT) allows the utilization of x-ray information as image priors for improving optical tomography reconstruction. To fully capitalize on this capacity, we consider a framework for the automatic and fast detection of different anatomic structures in murine XCT images. To accurately differentiate between different structures such as bone, lung, and heart, a combination of image processing steps including thresholding, seed growing, and signal detection are found to offer optimal segmentation performance. The algorithm and its utilization in an inverse FMT scheme that uses priors is demonstrated on mouse images.

Video Rate Optoacoustic Tomography of Mouse Kidney Perfusion

Optics Letters. Jul, 2010  |  Pubmed ID: 20634868

Optoacoustic tomography can visualize optical contrast in tissues while capitalizing on the advantages of ultrasound, such as high spatial resolution and fast imaging capabilities. We report a novel multispectral optoacoustic tomography system for deep tissue small animal imaging. The previously undocumented capacity of whole-body optoacoustic tomography at a video rate has been demonstrated by visualizing mouse kidney perfusion using Indocyanine Green in vivo.

Going Deeper Than Microscopy: the Optical Imaging Frontier in Biology

Nature Methods. Aug, 2010  |  Pubmed ID: 20676081

Optical microscopy has been a fundamental tool of biological discovery for more than three centuries, but its in vivo tissue imaging ability has been restricted by light scattering to superficial investigations, even when confocal or multiphoton methods are used. Recent advances in optical and optoacoustic (photoacoustic) imaging now allow imaging at depths and resolutions unprecedented for optical methods. These abilities are increasingly important to understand the dynamic interactions of cellular processes at different systems levels, a major challenge of postgenome biology. This Review discusses promising photonic methods that have the ability to visualize cellular and subcellular components in tissues across different penetration scales. The methods are classified into microscopic, mesoscopic and macroscopic approaches, according to the tissue depth at which they operate. Key characteristics associated with different imaging implementations are described and the potential of these technologies in biological applications is discussed.

Real-time Imaging of Cardiovascular Dynamics and Circulating Gold Nanorods with Multispectral Optoacoustic Tomography

Optics Express. Sep, 2010  |  Pubmed ID: 20940855

Macroscopic visualization of functional and molecular features of cardiovascular disease is emerging as an important tool in basic research and clinical translation of new diagnostic and therapeutic strategies. We showcase the application of Multispectral Optoacoustic Tomography (MSOT) techniques to noninvasively image different aspects of the mouse cardiovascular system macroscopically in real-time and in vivo, an unprecedented ability compared to optical or optoacoustic (photoacoustic) imaging approaches documented so far. In particular, we demonstrate imaging of the carotid arteries, the aorta and the cardiac wall. We further demonstrate the ability to dynamically visualize circulating gold nanorods that can be used to enhance contrast and be extended to molecular imaging applications. We discuss the potential of this imaging ability in cardiovascular disease (CVD) research and clinical applications.

Near-field Radiofrequency Thermoacoustic Tomography with Impulse Excitation

Medical Physics. Sep, 2010  |  Pubmed ID: 20964177

Imaging performance of radiofrequency and microwave-based thermoacoustic tomography systems is mainly determined by the ability to deposit a substantial amount of electromagnetic energy within ultrashort time duration. Pulses of nanosecond-range duration that can carry hundreds of millijoules energy are ideal for obtaining good signal-to-noise and spatial resolution in many biological imaging applications. However, existing implementations are based on modulated-carrier-frequency amplification solutions, which are generally costly and cannot achieve ultrahigh-peak-power requirements essential for optimal thermoacoustic signal generation.

Optoacoustic Tomography with Varying Illumination and Non-uniform Detection Patterns

Journal of the Optical Society of America. A, Optics, Image Science, and Vision. Nov, 2010  |  Pubmed ID: 21045914

Quantification of tissue morphology and biomarker distribution by means of optoacoustic tomography is an important and longstanding challenge, mainly caused by the complex heterogeneous structure of biological tissues as well as the lack of accurate and robust reconstruction algorithms. The recently introduced model-based inversion approaches were shown to mitigate some of reconstruction artifacts associated with the commonly used back-projection schemes, while providing an excellent platform for obtaining quantified maps of optical energy deposition in experimental configurations of various complexity. In this work, we introduce a weighted model-based approach, capable of overcoming reconstruction challenges caused by per-projection variations of object's illumination and other partial illumination effects. The universal weighting procedure is equally shown to reduce reconstruction artifacts associated with other experimental imperfections, such as non-uniform transducer sensitivity fields. Significant improvements in image fidelity and quantification are showcased both numerically and experimentally on tissue phantoms and mice.

Performance Evaluation of Adaptive Meshing Algorithms for Fluorescence Diffuse Optical Tomography Using Experimental Data

Optics Letters. Nov, 2010  |  Pubmed ID: 21081977

Fluorescence diffuse optical tomography (FDOT) is a computationally demanding imaging problem. The discretizations of FDOT forward and inverse problems pose a trade-off between the accuracy and the computational efficiency of the image reconstruction. To address this trade-off, we analyzed the effect of discretization on the accuracy of FDOT imaging and proposed novel adaptive meshing algorithms for FDOT in a series of studies. In this Letter, we apply these new adaptive meshing algorithms to FDOT imaging using real data from a phantom experiment to demonstrate the practical advantages of our algorithms in FDOT image reconstruction.

Current Concepts and Future Perspectives on Surgical Optical Imaging in Cancer

Journal of Biomedical Optics. Nov-Dec, 2010  |  Pubmed ID: 21198198

There are vibrant developments of optical imaging systems and contrast-enhancing methods that are geared to enhancing surgical vision and the outcome of surgical procedures. Such optical technologies designed for intraoperative use can offer high integration in the operating room compared to conventional radiological modalities adapted to intraoperative applications. Simple fluorescence epi-illumination imaging, in particular, appears attractive but may lead to inaccurate observations due to the complex nature of photon-tissue interaction. Of importance therefore are emerging methods that account for the background optical property variation in tissues and can offer accurate, quantitative imaging that eliminates the appearance of false negatives or positives. In parallel, other nonfluorescent optical imaging methods are summarized and overall progress in surgical optical imaging applications is outlined. Key future directions that have the potential to shift the paradigm of surgical health care are also discussed.

Data Specific Spatially Varying Regularization for Multimodal Fluorescence Molecular Tomography

IEEE Transactions on Medical Imaging. Feb, 2010  |  Pubmed ID: 19758858

Fluorescence molecular tomography (FMT) allows in vivo localization and quantification of fluorescence biodistributions in whole animals. The ill-posed nature of the tomographic reconstruction problem, however, limits the attainable resolution. Improvements in resolution and overall imaging performance can be achieved by forming image priors from geometric information obtained by a secondary anatomical or functional high-resolution imaging modality such as X-ray computed tomography or magnetic resonance imaging. A particular challenge in using image priors is to avoid the use of assumptions that may bias the solution and reduced the accuracy of the inverse problem. This is particularly relevant in FMT inversions where there is not an evident link between secondary geometric information and the underlying fluorescence biodistribution. We present here a new, two step approach to incorporating structural priors into the FMT inverse problem. By using the anatomic information to define a low dimensional inverse problem, we obtain a solution which we then use to determine the parameters defining a spatially varying regularization matrix for the full resolution problem. The regularization term is thus customized for each data set and is guided by the data rather than depending only on user defined a priori assumptions. Results are presented for both simulated and experimental data sets, and show significant improvements in image quality as compared to traditional regularization techniques.

Hybrid System for Simultaneous Fluorescence and X-ray Computed Tomography

IEEE Transactions on Medical Imaging. Feb, 2010  |  Pubmed ID: 19906585

A hybrid imaging system for simultaneous fluorescence tomography and X-ray computed tomography (XCT) of small animals has been developed and presented. The system capitalizes on the imaging power of a 360 ( degrees )-projection free-space fluorescence tomography system, implemented within a microcomputed tomography scanner. Image acquisition is based on techniques that automatically adjust a series of imaging parameters to offer a high dynamic range dataset. Image segmentation further allows the incorporation of structural priors in the optical reconstruction problem to improve the imaging performance. The functional system characteristics are showcased, and images from a brain imaging study are shown, which are reconstructed using XCT-derived priors into the optical forward problem.

Indocyanine Green Enables Near-infrared Fluorescence Imaging of Lipid-rich, Inflamed Atherosclerotic Plaques

Science Translational Medicine. May, 2011  |  Pubmed ID: 21613624

New high-resolution molecular and structural imaging strategies are needed to visualize high-risk plaques that are likely to cause acute myocardial infarction, because current diagnostic methods do not reliably identify at-risk subjects. Although molecular imaging agents are available for low-resolution detection of atherosclerosis in large arteries, a lack of imaging agents coupled to high-resolution modalities has limited molecular imaging of atherosclerosis in the smaller coronary arteries. Here, we have demonstrated that indocyanine green (ICG), a Food and Drug Administration-approved near-infrared fluorescence (NIRF)-emitting compound, targets atheromas within 20 min of injection and provides sufficient signal enhancement for in vivo detection of lipid-rich, inflamed, coronary-sized plaques in atherosclerotic rabbits. In vivo NIRF sensing was achieved with an intravascular wire in the aorta, a vessel of comparable caliber to human coronary arteries. Ex vivo fluorescence reflectance imaging showed high plaque target-to-background ratios in atheroma-bearing rabbits injected with ICG compared to atheroma-bearing rabbits injected with saline. In vitro studies using human macrophages established that ICG preferentially targets lipid-loaded macrophages. In an early clinical study of human atheroma specimens from four patients, we found that ICG colocalized with plaque macrophages and lipids. The atheroma-targeting capability of ICG has the potential to accelerate the clinical development of NIRF molecular imaging of high-risk plaques in humans.

Compressive Sensing for Biomedical Imaging

IEEE Transactions on Medical Imaging. May, 2011  |  Pubmed ID: 21692237

Detection of Irradiation-induced, Membrane Heat Shock Protein 70 (Hsp70) in Mouse Tumors Using Hsp70 Fab Fragment

Radiotherapy and Oncology : Journal of the European Society for Therapeutic Radiology and Oncology. Jun, 2011  |  Pubmed ID: 21704400

The major stress-inducible heat shock protein 70 (Hsp70) is frequently overexpressed in highly aggressive tumors, and elevated intracellular Hsp70 levels mediate protection against apoptosis. Following therapeutic intervention, such as ionizing irradiation, translocation of cytosolic Hsp70 to the plasma membrane is selectively increased in tumor cells and therefore, membrane Hsp70 might serve as a therapy-inducible, tumor-specific target structure.

Multispectral Optoacoustic Tomography of Matrix Metalloproteinase Activity in Vulnerable Human Carotid Plaques

Molecular Imaging and Biology : MIB : the Official Publication of the Academy of Molecular Imaging. Jul, 2011  |  Pubmed ID: 21720908

AIMS: Elevated expression of cathepsins, integrins and matrix metalloproteinases (MMPs) is typically associated with atherosclerotic plaque instability. While fluorescent tagging of such molecules has been amply demonstrated, no imaging method was so far shown capable of resolving these inflammation-associated tags with high fidelity and resolution beyond microscopic depths. This study is aimed at demonstrating a new method with high potential for noninvasive clinical cardiovascular diagnostics of vulnerable plaques using high-resolution deep-tissue multispectral optoacoustic tomography (MSOT) technology. METHODS AND RESULTS: MMP-sensitive activatable fluorescent probe (MMPSense™ 680) was applied to human carotid plaques from symptomatic patients. Atherosclerotic activity was detected by tuning MSOT wavelengths to activation-dependent absorption changes of the molecules, structurally modified in the presence of enzymes. MSOT analysis simultaneously provided morphology along with heterogeneous MMP activity with better than 200 micron resolution throughout the intact plaque tissue. The results corresponded well with epi-fluorescence images made from thin cryosections. Elevated MMP activity was further confirmed by in situ zymography, accompanied by increased macrophage influx. CONCLUSIONS: We demonstrated, for the first time to our knowledge, the ability of MSOT to provide volumetric images of activatable molecular probe distribution deep within optically diffuse tissues. High-resolution mapping of MMP activity was achieved deep in the vulnerable plaque of intact human carotid specimens. This performance directly relates to pre-clinical screening applications in animal models and to clinical decision potential as it might eventually allow for highly specific visualization and staging of plaque vulnerability thus impacting therapeutic clinical decision making.

Volumetric Real-time Multispectral Optoacoustic Tomography of Biomarkers

Nature Protocols. Aug, 2011  |  Pubmed ID: 21738125

Multispectral optoacoustic tomography (MSOT) has recently been developed to enable visualization of optical contrast and tissue biomarkers, with resolution and speed representative of ultrasound. In the implementation described here, MSOT enables operation in real-time mode by capturing single cross-sectional images in <1 ms from living small animals (e.g., mice) and other tissues of similar dimensions. At the core of the method is illumination of the object using multiple wavelengths in order to resolve spectrally distinct biomarkers over background tissue chromophores. The system allows horizontal placement of a mouse in the imaging chamber and three-dimensional scanning of the entire body without the need to immerse the mouse in water. Here we provide a detailed description of the MSOT scanner components, system calibration, selection of image reconstruction algorithms and animal handling. Overall, the entire protocol can be completed within 15-30 min for acquisition of a whole-body multispectral data set from a living mouse.

Model-based Optoacoustic Inversion with Arbitrary-shape Detectors

Medical Physics. Jul, 2011  |  Pubmed ID: 21859030

Optoacoustic imaging enables mapping the optical absorption of biological tissue using optical excitation and acoustic detection. Although most image-reconstruction algorithms are based on the assumption of a detector with an isotropic sensitivity, the geometry of the detector often leads to a response with spatially dependent magnitude and bandwidth. This effect may lead to attenuation or distortion in the recorded signal and, consequently, in the reconstructed image.

The Effects of Acoustic Attenuation in Optoacoustic Signals

Physics in Medicine and Biology. Sep, 2011  |  Pubmed ID: 21873768

In this paper, it is demonstrated that the effects of acoustic attenuation may play a significant role in establishing the quality of tomographic optoacoustic reconstructions. Accordingly, spatially dependent reduction of signal amplitude leads to quantification errors in the reconstructed distribution of the optical absorption coefficient while signal broadening causes loss of image resolution. Here we propose a correction algorithm for accounting for attenuation effects, which is applicable in both the time and frequency domains. It is further investigated which part of the optoacoustic signal spectrum is practically affected by those effects in realistic imaging scenarios. The validity and benefits of the suggested modelling and correction approaches are experimentally validated in phantom measurements.

Intraoperative Tumor-specific Fluorescence Imaging in Ovarian Cancer by Folate Receptor-α Targeting: First In-human Results

Nature Medicine. Oct, 2011  |  Pubmed ID: 21926976

The prognosis in advanced-stage ovarian cancer remains poor. Tumor-specific intraoperative fluorescence imaging may improve staging and debulking efforts in cytoreductive surgery and thereby improve prognosis. The overexpression of folate receptor-α (FR-α) in 90-95% of epithelial ovarian cancers prompted the investigation of intraoperative tumor-specific fluorescence imaging in ovarian cancer surgery using an FR-α-targeted fluorescent agent. In patients with ovarian cancer, intraoperative tumor-specific fluorescence imaging with an FR-α-targeted fluorescent agent showcased the potential applications in patients with ovarian cancer for improved intraoperative staging and more radical cytoreductive surgery.

Intraoperative Near-infrared Fluorescence Tumor Imaging with Vascular Endothelial Growth Factor and Human Epidermal Growth Factor Receptor 2 Targeting Antibodies

Journal of Nuclear Medicine : Official Publication, Society of Nuclear Medicine. Nov, 2011  |  Pubmed ID: 21990576

Fluorescence imaging is currently attracting much interest as a method for intraoperative tumor detection, but most current tracers lack tumor specificity. Therefore, this technique can be further improved by tumor-specific detection. With tumor-targeted antibodies bound to a radioactive label, tumor-specific SPECT or PET is feasible in the clinical setting. The aim of the present study was to apply antibody-based tumor detection to intraoperative optical imaging, using preclinical in vivo mouse models.

Clinical Translation of Optical and Optoacoustic Imaging

Philosophical Transactions. Series A, Mathematical, Physical, and Engineering Sciences. Nov, 2011  |  Pubmed ID: 22006913

Macroscopic optical imaging has rather humble technical origins; it has been mostly implemented by photographic means using appropriate filters, a light source and a camera yielding images of tissues. This approach relates to human vision and perception, and is simple to implement and use. Therefore, it has found wide acceptance, especially in recording fluorescence and bioluminescence signals. Yet, the difficulty in resolving depth and the dependence of the light intensity recorded on tissue optical properties may compromise the accuracy of the approach. Recently, optical technology has seen significant advances that bring a new performance level in optical investigations. Quantitative real-time multi-spectral optical and optoacoustic (photoacoustic) methods enable high-resolution quantitative imaging of tissue and disease biomarkers and can significantly enhance medical vision in diagnostic or interventional procedures such as dermatology, endoscopy, surgery, and various vascular and intravascular imaging applications. This performance is showcased herein and examples are given to illustrate how it is possible to shift the paradigm of optical clinical translation.

Optoacoustic Imaging: an Emerging Modality for the Gastrointestinal Tract

Gastroenterology. Dec, 2011  |  Pubmed ID: 22015916

Multispectral Optoacoustic Tomography by Means of Normalized Spectral Ratio

Optics Letters. Nov, 2011  |  Pubmed ID: 22048356

Quantification of biomarkers using multispectral optoacoustic tomography can be challenging due to photon fluence variations with depth and spatially heterogeneous tissue optical properties. Herein we introduce a spectral ratio approach that accounts for photon fluence variations. The performance and imaging improvement achieved with the proposed method is showcased both numerically and experimentally in phantoms and mice.

Intra-arterial Catheter for Simultaneous Microstructural and Molecular Imaging in Vivo

Nature Medicine. Dec, 2011  |  Pubmed ID: 22057345

Advancing understanding of human coronary artery disease requires new methods that can be used in patients for studying atherosclerotic plaque microstructure in relation to the molecular mechanisms that underlie its initiation, progression and clinical complications, including myocardial infarction and sudden cardiac death. Here we report a dual-modality intra-arterial catheter for simultaneous microstructural and molecular imaging in vivo using a combination of optical frequency domain imaging (OFDI) and near-infrared fluorescence (NIRF) imaging. By providing simultaneous molecular information in the context of the surrounding tissue microstructure, this new catheter could provide new opportunities for investigating coronary atherosclerosis and stent healing and for identifying high-risk biological and structural coronary arterial plaques in vivo.

Revisiting the Normalized Born Approximation: Effects of Scattering

Optics Letters. Nov, 2011  |  Pubmed ID: 22089553

The normalized Born approximation has been suggested as a ratiometric method in fluorescence molecular tomography (FMT) applications, to account for heterogeneity variations. The method enabled practical inversions, as it offered fluorescence reconstruction accuracy over a wide range of absorption heterogeneity, while also accounting for unknown experimental factors, such as the various system gains and losses. Yet it was noted that scattering variations affect the robustness and accuracy. Herein we decompose the effects of absorption and scattering and capitalize on the recent development of hybrid FMT/x-ray computed tomography imaging methods to proposed amendments to the method, which improve the overall accuracy of the approach.

Intraoperative Multispectral Fluorescence Imaging for the Detection of the Sentinel Lymph Node in Cervical Cancer: a Novel Concept

Molecular Imaging and Biology : MIB : the Official Publication of the Academy of Molecular Imaging. Oct, 2011  |  Pubmed ID: 20835767

Real-time intraoperative near-infrared fluorescence (NIRF) imaging is a promising technique for lymphatic mapping and sentinel lymph node (SLN) detection. The purpose of this technical feasibility pilot study was to evaluate the applicability of NIRF imaging with indocyanin green (ICG) for the detection of the SLN in cervical cancer.

Imaging the Bio-distribution of Fluorescent Probes Using Multispectral Epi-illumination Cryoslicing Imaging

Molecular Imaging and Biology : MIB : the Official Publication of the Academy of Molecular Imaging. Oct, 2011  |  Pubmed ID: 20838910

The increasing availability of fluorescent probes for in vivo optical imaging enables the interrogation of complex biological processes in small animals serving as models for human-like tissue function and disease. However, the validation of probe bio-distribution during their development or the study of different disease models, in support of in vivo imaging studies, is not straightforward.

Statistical Approach for Optoacoustic Image Reconstruction in the Presence of Strong Acoustic Heterogeneities

IEEE Transactions on Medical Imaging. Feb, 2011  |  Pubmed ID: 20876007

A method is presented to reduce artefacts produced in optoacoustic tomography images due to internal reflection or scattering of the acoustic waves. It is based on weighting the tomographic contribution of each detector with the probability that a signal affected by acoustic mismatches is measured at that position. The correction method does not require a priori knowledge of the acoustic or optical properties of the imaged sample. Performance tests were made with agar phantoms that included air gaps for mimicking strong acoustic reflections as well as with an acoustically heterogeneous adult Zebrafish. The results obtained with the method proposed show a clear reduction of the artefacts with respect to the original images reconstructed with filtered back-projection algorithm. This performance is directly related to in vivo small animal imaging applications involving imaging in the presence of bones, lungs, and other highly mismatched organs.

In Vivo Imaging of CT26 Mouse Tumours by Using CmHsp70.1 Monoclonal Antibody

Journal of Cellular and Molecular Medicine. Apr, 2011  |  Pubmed ID: 20406322

The major stress-inducible heat shock protein 70 (Hsp70) is frequently present on the cell surface of human tumours, but not on normal cells. Herein, the binding characteristics of the cmHsp70.1 mouse monoclonal antibody (mAb) were evaluated in vitro and in a syngeneic tumour mouse model. More than 50% of the CT26 mouse colon carcinoma cells express Hsp70 on their cell surface at 4°C. After a temperature shift to 37°C, the cmHsp70.1-fluorescein isothiocyanate mAb translocates into early endosomes and lysosomes. Intraoperative and near-infrared fluorescence imaging revealed an enrichment of Cy5.5-conjugated mAb cmHsp70.1, but not an identically labelled IgG1 isotype-matched control, in i.p. and s.c. located CT26 tumours, as soon as 30 min. after i.v. injection into the tail vein. Due to the rapid turnover rate of membrane-bound Hsp70, the fluorescence-labelled cmHsp70.1 mAb became endocytosed and accumulated in the tumour, reaching a maximum after 24 hrs and remained detectable at least up to 96 hrs after a single i.v. injection. The tumour-selective internalization of mAb cmHsp70.1 at the physiological temperature of 37°C might enable a targeted uptake of toxins or radionuclides into Hsp70 membrane-positive tumours. The anti-tumoral activity of the cmHsp70.1 mAb is further supported by its capacity to mediate antibody-dependent cytotoxicity.

Progress on Multimodal Molecular / Anatomical Intravascular Imaging of Coronary Vessels Combining Near Infrared Fluorescence and Ultrasound

Conference Proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference. Aug, 2011  |  Pubmed ID: 22254510

The use of intravascular imaging modalities for the detection and assessment of atherosclerotic plaque is becoming increasingly useful. Current clinical invasive modalities assess the presence of plaque using anatomical information and include Intravascular Ultrasound (IVUS) and Optical Coherence Tomography (OCT). However, such modalities cannot take into account underlying functional biological information, which can however be revealed with the use of molecular imaging. Consequently, intravascular molecular imaging is emerging as a powerful approach. We have developed such a Near-Infrared Fluorescence (NIRF) imaging system and showcased, in both phantom and in-vivo (rabbit) experiments, its potential to successfully detect inflamed atherosclerotic plaques, using appropriate fluorescent probes. Here, we discuss some limitations of the current system and suggest the combined use of the NIRF and IVUS imaging systems as a means for more accurate assessment of atherosclerotic plaque. We include some results and models that showcase the potential power of this kind of hybrid imaging.

Optoacoustic Methods for Frequency Calibration of Ultrasonic Sensors

IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control. Feb, 2011  |  Pubmed ID: 21342817

The frequency response of ultrasonic detectors is commonly calibrated by finding their sensitivity to incident plane waves at discrete frequencies. For certain applications, such as the emerging field of optoacoustic tomography, it is the response to point sources emitting broadband spectra that needs to be found instead. Although these two distinct sensitivity characteristics are interchangeable in the case of a flat detector and a point source at infinity, it is not the case for detectors with size considerably larger than the acoustic wavelength of interest or those having a focused aperture. Such geometries, which are common in optoacoustics, require direct calibration of the acoustic detector using a point source placed in the relevant position. In this paper, we report on novel cross-validating optoacoustic methods for measuring the frequency response of wideband acoustic sensors. The approach developed does not require pre-calibrated hydrophones and therefore can be readily adopted in any existing optoacoustic measurement configuration. The methods are successfully confirmed experimentally by measuring the frequency response of a common piezoelectric detector having a cylindrically focused shape.

Blind Source Unmixing in Multi-spectral Optoacoustic Tomography

Optics Express. Feb, 2011  |  Pubmed ID: 21369139

Multispectral optoacoustic (photoacoustic) tomography (MSOT) is a hybrid modality that can image through several millimeters to centimeters of diffuse tissues, attaining resolutions typical of ultrasound imaging. The method can further identify tissue biomarkers by decomposing the spectral contributions of different photo-absorbing molecules of interest. In this work we investigate the performance of blind source unmixing methods and spectral fitting approaches in decomposing the contributions of fluorescent dyes from the tissue background, based on MSOT measurements in mice. We find blind unmixing as a promising method for accurate MSOT decomposition, suitable also for spectral unmixing in fluorescence imaging. We further demonstrate its capacity with temporal unmixing on real-time MSOT data obtained in-vivo for enhancing the visualization of absorber agent flow in the mouse vascular system.

In Vivo Tomographic Imaging of Red-shifted Fluorescent Proteins

Biomedical Optics Express. 2011  |  Pubmed ID: 21483611

We have developed a spectral inversion method for three-dimensional tomography of far-red and near-infrared fluorescent proteins in animals. The method was developed in particular to address the steep light absorption transition of hemoglobin from the visible to the far-red occurring around 600 nm. Using an orthotopic mouse model of brain tumors expressing the red-shifted fluorescent protein mCherry, we demonstrate significant improvements in imaging accuracy over single-wavelength whole body reconstructions. Furthermore, we show an improvement in sensitivity of at least an order of magnitude over green fluorescent protein (GFP) for whole body imaging. We discuss how additional sensitivity gains are expected with the use of further red-shifted fluorescent proteins and we explain the differences and potential advantages of this approach over two-dimensional planar imaging methods.

Enhancing Surgical Vision by Using Real-time Imaging of αvβ3-integrin Targeted Near-infrared Fluorescent Agent

Annals of Surgical Oncology. Nov, 2011  |  Pubmed ID: 21509632

This study was designed to improve the surgical procedure and outcome of cancer surgery by means of real-time molecular imaging feedback of tumor spread and margin delineation using targeted near-infrared fluorescent probes with specificity to tumor biomarkers. Surgical excision of cancer often is confronted with difficulties in the identification of cancer spread and the accurate delineation of tumor margins. Currently, the assessment of tumor borders is afforded by postoperative pathology or, less reliably, intraoperative frozen sectioning. Fluorescence imaging is a natural modality for intraoperative use by directly relating to the surgeon's vision and offers highly attractive characteristics, such as high-resolution, sensitivity, and portability. Via the use of targeted probes it also becomes highly tumor-specific and can lead to significant improvements in surgical procedures and outcome.

Model-based Optoacoustic Inversions with Incomplete Projection Data

Medical Physics. Mar, 2011  |  Pubmed ID: 21520882

Optoacoustic imaging is an emerging noninvasive imaging modality that can resolve optical contrast through several millimeters to centimeters of tissue with diffraction-limited resolution of ultrasound. Yet, quantified reconstruction of tissue absorption maps requires optoacoustic signals to be collected from as many locations around the object as possible. In many tomographic imaging scenarios, however, only limited-view or partial projection data are available, which has been shown to generate image artifacts and overall loss of quantification accuracy.

Reconstruction of Fluorescence Distribution Hidden in Biological Tissue Using Mesoscopic Epifluorescence Tomography

Journal of Biomedical Optics. Apr, 2011  |  Pubmed ID: 21529074

Mesoscopic epifluorescence tomography is a novel technique that discovers fluorescence bio-distribution in small animals by tomographic means in reflectance geometry. A collimated laser beam is scanned over the skin surface to excite fluorophores hidden within the tissue while a CCD camera acquires an image of the fluorescence emission for each source position. This configuration is highly efficient in the visible spectrum range where trans-illumination imaging of small animals is not feasible due to the high tissue absorption and scattering in biological organisms. The reconstruction algorithm is similar to the one used in fluorescence molecular tomography. However, diffusion theory cannot be employed since the source-detector separation for most image pixels is comparable to or below the scattering length of the tissue. Instead Monte Carlo simulations are utilized to predict the sensitivity functions. In a phantom study we show the effect of using enhanced source grid arrangements during the data acquisition and the reconstruction process to minimize boundary artefacts. Furthermore, we present ex vivo data that show high spatial resolution and quantitative accuracy in heterogeneous tissues using GFP-like fluorescence in B6-albino mice up to a depth of 1100 μm.

Near-field Thermoacoustic Tomography of Small Animals

Physics in Medicine and Biology. Jun, 2011  |  Pubmed ID: 21572232

Near-field radiofrequency thermoacoustic (NRT) tomography is a new imaging method that was developed to mitigate limitations of conventional thermoacoustic imaging approaches, related to hard compromises between signal strength and spatial resolution. By utilizing ultrahigh-energy electromagnetic impulses at ∼20 ns duration along with improved energy absorption coupling in the near-field, this method can deliver high-resolution images without compromising signal to noise ratio. NRT is a promising modality, offering cost-effectiveness and ease of implementation and it can be conveniently scaled to image small animals and humans. However, several of the performance metrics of the method are not yet documented. In this paper, we characterize the expected imaging performance via numerical simulations based on a finite-integration time-domain (FITD) technique and experiments using tissue mimicking phantoms and different biological samples. Furthermore, we show for the first time whole-body tomographic imaging results from mice, revealing clear anatomical details along with highly dissipative inclusions introduced for control. The best spatial resolution achieved for those experiments was 150 µm.

High-sensitivity Compact Ultrasonic Detector Based on a Pi-phase-shifted Fiber Bragg Grating

Optics Letters. May, 2011  |  Pubmed ID: 21593906

A highly sensitive compact hydrophone, based on a pi-phase-shifted fiber Bragg grating, has been developed for the measurement of wideband ultrasonic fields. The grating exhibits a sharp resonance, whose centroid wavelength is pressure sensitive. The resonance is monitored by a continuous-wave (CW) laser to measure ultrasound-induced pressure variations within the grating. In contrast to standard fiber sensors, the high finesse of the resonance--which is the reason for the sensor's high sensitivity--is not associated with a long propagation length. Light localization around the phase shift reduces the effective size of the sensor below that of the grating and is scaled inversely with the resonance spectral width. In our system, an effective sensor length of 270 μm, pressure sensitivity of 440 Pa, and effective bandwidth of 10 MHz were achieved. This performance makes our design attractive for medical imaging applications, such as optoacoustic tomography, in which compact, sensitive, and wideband acoustic detectors are required.

Two-dimensional Intravascular Near-infrared Fluorescence Molecular Imaging of Inflammation in Atherosclerosis and Stent-induced Vascular Injury

Journal of the American College of Cardiology. Jun, 2011  |  Pubmed ID: 21679853

This study sought to develop a 2-dimensional (2D) intravascular near-infrared fluorescence (NIRF) imaging strategy for investigation of arterial inflammation in coronary-sized vessels.

Fast Multispectral Optoacoustic Tomography (MSOT) for Dynamic Imaging of Pharmacokinetics and Biodistribution in Multiple Organs

PloS One. 2012  |  Pubmed ID: 22295087

The characterization of pharmacokinetic and biodistribution profiles is an essential step in the development process of new candidate drugs or imaging agents. Simultaneously, the assessment of organ function related to the uptake and clearance of drugs is of great importance. To this end, we demonstrate an imaging platform capable of high-rate characterization of the dynamics of fluorescent agents in multiple organs using multispectral optoacoustic tomography (MSOT). A spatial resolution of approximately 150 µm through mouse cross-sections allowed us to image blood vessels, the kidneys, the liver and the gall bladder. In particular, MSOT was employed to characterize the removal of indocyanine green from the systemic circulation and its time-resolved uptake in the liver and gallbladder. Furthermore, it was possible to track the uptake of a carboxylate dye in separate regions of the kidneys. The results demonstrate the acquisition of agent concentration metrics at rates of 10 samples per second at a single wavelength and 17 s per multispectral sample with 10 signal averages at each of 5 wavelengths. Overall, such imaging performance introduces previously undocumented capabilities of fast, high resolution in vivo imaging of the fate of optical agents for drug discovery and basic biological research.

Opto-acoustic Imaging of Drug Discovery Biomarkers

Current Pharmaceutical Biotechnology. Feb, 2012  |  Pubmed ID: 22335481

Optical imaging has seen significant developments over the past decade as an investigational tool for in-vivo visualization of cellular and sub-cellular events. With the recent addition of optoacoustic (photoacoustic) methods, in particular multi-spectral opto-acoustictomography (MSOT), to the already rich armamentarium of photonic methods the capacity of optical molecular imaging across scales haswidened significantly. MSOT brings unique features into optical imaging, namely high resolution optical imaging over several millimeters to centimeters of tissue depth and the ability to simultaneously resolve multiple tissue molecules and extrinsically administered optical or optoacoustic agents with physiological or molecular specificity. Here, we discuss the implications of utilizing MSOT in the context of drug discovery and review suitable optoacousticagents against disease and drug efficacy biomarkers. The combination of existing knowledge on generating optical targeted contrast, with the high resolution deep tissue visualization offered by MSOT, allows for the development of next-generation biological optical imaging and corresponding drug discovery applications.

Deep Tissue Optical and Optoacoustic Molecular Imaging Technologies for Small Animal Research and Drug Discovery

Current Pharmaceutical Biotechnology. Jan, 2012  |  Pubmed ID: 22216767

For centuries, biological discoveries were based on optical imaging, in particular microscopy but also several chromophoric assays and photographic approaches. With the recent emergence of methods appropriate for bio-marker in vivo staining, such as bioluminescence, fluorescent molecular probes and proteins, as well as nanoparticle-based targeted agents, significant attention has been shifted toward in vivo interrogations of different dynamic biological processes at the molecular level. This progress has been largely supported by the development of advanced tomographic imaging technologies suitable for obtaining volumetric visualization of bio-marker distributions in small animals at a whole-body or whole-organ scale, an imaging frontier that is not accessible by the existing tissue-sectioning microscopic techniques due to intensive light scattering beyond the depth of a few hundred microns. Major examples of such recently developed optical imaging modalities are reviewed here, including bioluminescence tomography (BLT), fluorescence molecular tomography (FMT), and optical projection tomography (OPT). The pharmaceutical imaging community has quickly appropriated itself of these novel forms of optical imaging, since they come with very compelling advantages, such as quantitative three-dimensional capabilities, direct correlation to the biological cultures, easiness and cost-effectiveness of use, and the use of safe non-ionizing radiation. Some multi-modality approaches, combining light with other imaging modalities such as X-Ray CT or MRI, giving the ability to acquire both an optical contrast reconstruction along with a hi-fidelity anatomical images, are also reviewed. A separate section is devoted to the hybrid imaging techniques based on the optoacoustic phenomenon, such as multispectral optoacoustic tomography (MSOT), which are poised to leverage the traditional contrast and specificity advantages of optical spectrum by delivering an ever powerful set of capabilities, including real-time operation and high spatial resolution, not affected by the scattering nature of biological tissues.

Optical and Opto-Acoustic Interventional Imaging

Annals of Biomedical Engineering. Jan, 2012  |  Pubmed ID: 22227974

Many clinical interventional procedures, such as surgery or endoscopy, are today still guided by human vision and perception. Human vision however is not sensitive or accurate in detecting a large range of disease biomarkers, for example cellular or molecular processes characteristic of disease. For this reason advanced optical and opto-acoustic (photo-acoustic) methods are considered for enabling a more versatile, sensitive and accurate detection of disease biomarkers and complement human vision in clinical decision making during interventions. Herein, we outline developments in emerging fluorescence and opto-acoustic sensing and imaging techniques that can lead to practical implementations toward improving interventional vision.