Two-dimensional Depth-resolved Mueller Matrix of Biological Tissue Measured with Double-beam Polarization-sensitive Optical Coherence Tomography

Optics Letters. Jan, 2002  |  Pubmed ID: 18007725

A double-beam polarization-sensitive system based on optical coherence tomography was built to measure the Mueller matrix of scattering biological tissue with high spatial resolution. The Jones matrix of a sample can be determined with a single scan and subsequently converted into an equivalent nondepolarizing Mueller matrix. As a result, the system can be used to measure the Mueller matrix of an unstable sample, such as soft tissue. The polarization parameters of a porcine tendon, including magnitude and orientation of birefringence and diattenuation, were extracted by decomposition of the measured Mueller matrix.

Ultrasound-mediated Biophotonic Imaging: a Review of Acousto-optical Tomography and Photo-acoustic Tomography

Disease Markers. -2004, 2003  |  Pubmed ID: 15096709

This article reviews two types of ultrasound-mediated biophotonic imaging-acousto-optical tomography (AOT, also called ultrasound-modulated optical tomography) and photo-acoustic tomography (PAT, also called opto-acoustic or thermo-acoustic tomography)-both of which are based on non-ionizing optical and ultrasonic waves. The goal of these technologies is to combine the contrast advantage of the optical properties and the resolution advantage of ultrasound. In these two technologies, the imaging contrast is based primarily on the optical properties of biological tissues, and the imaging resolution is based primarily on the ultrasonic waves that either are provided externally or produced internally, within the biological tissues. In fact, ultrasonic mediation overcomes both the resolution disadvantage of pure optical imaging in thick tissues and the contrast and speckle disadvantages of pure ultrasonic imaging. In our discussion of AOT, the relationship between modulation depth and acoustic amplitude is clarified. Potential clinical applications of ultrasound-mediated biophotonic imaging include early cancer detection, functional imaging, and molecular imaging.

Analytic Explanation of Spatial Resolution Related to Bandwidth and Detector Aperture Size in Thermoacoustic or Photoacoustic Reconstruction

Physical Review. E, Statistical, Nonlinear, and Soft Matter Physics. May, 2003  |  Pubmed ID: 12786294

An analytic explanation of the spatial resolution in thermoacoustic or photoacoustic reconstruction is presented. Three types of specific recording geometries, including spherical, planar, and cylindrical surface, as well as other general cases, are investigated. Analytic expressions of the point-spread functions (PSF's), as a function of the bandwidth of the measurement system and the finite size of the detector aperture, are derived based on rigorous reconstruction formulas. The analyses clearly reveal that the dependence of the PSF's on the bandwidth of all recording geometries shares the same space-invariant expression while the dependence on the aperture size of the detector differs. The bandwidth affects both axial and lateral resolutions; in contrast, the detector aperture blurs the lateral resolution greatly but the axial resolution only slightly.

Noninvasive Laser-induced Photoacoustic Tomography for Structural and Functional in Vivo Imaging of the Brain

Nature Biotechnology. Jul, 2003  |  Pubmed ID: 12808463

Imaging techniques based on optical contrast analysis can be used to visualize dynamic and functional properties of the nervous system via optical signals resulting from changes in blood volume, oxygen consumption and cellular swelling associated with brain physiology and pathology. Here we report in vivo noninvasive transdermal and transcranial imaging of the structure and function of rat brains by means of laser-induced photoacoustic tomography (PAT). The advantage of PAT over pure optical imaging is that it retains intrinsic optical contrast characteristics while taking advantage of the diffraction-limited high spatial resolution of ultrasound. We accurately mapped rat brain structures, with and without lesions, and functional cerebral hemodynamic changes in cortical blood vessels around the whisker-barrel cortex in response to whisker stimulation. We also imaged hyperoxia- and hypoxia-induced cerebral hemodynamic changes. This neuroimaging modality holds promise for applications in neurophysiology, neuropathology and neurotherapy.

Transmission- and Side-detection Configurations in Ultrasound-modulated Optical Tomography of Thick Biological Tissues

Applied Optics. Jul, 2003  |  Pubmed ID: 12868851

Ultrasound-modulated optical tomography of thick biological tissues was studied based on speckle-contrast detection. Speckle decorrelation was investigated with biological tissue samples of various thicknesses. Images of optically absorbing objects buried in biological tissue samples with thicknesses up to 50 mm were obtained in a transmission-detection configuration. The image contrast was more than 30%, and the spatial resolution was approximately 2 mm. In addition, a side-detection scheme along with two specific configurations were examined, and the advantages were demonstrated. Experimental results implied feasibility of applying the ultrasound-modulation technique to characterize optical properties in inhomogeneous biological tissues.

Optical-fiber-based Mueller Optical Coherence Tomography

Optics Letters. Jul, 2003  |  Pubmed ID: 12885022

An optical-fiber-based multichannel polarization-sensitive Mueller optical coherence tomography (OCT) system was built to acquire the Jones or Mueller matrix of a scattering medium, such as biological tissue. For the first time to our knowledge, fiber-based polarization-sensitive OCT was dynamically calibrated to eliminate the polarization distortion caused by the single-mode optical fiber in the sample arm, thereby overcoming a key technical impediment to the application of optical fibers in this technology. The round-trip Jones matrix of the sampling fiber was acquired from the reflecting surface of the sample for each depth scan (A scan) with our OCT system. A new rigorous algorithm was then used to retrieve the calibrated polarization properties of the sample. This algorithm was validated with experimental data. The skin of a rat was imaged with this fiber-based system.

Time-domain Reconstruction Algorithms and Numerical Simulations for Thermoacoustic Tomography in Various Geometries

IEEE Transactions on Bio-medical Engineering. Sep, 2003  |  Pubmed ID: 12943276

In this paper, we present time-domain reconstruction algorithms for the thermoacoustic imaging of biological tissues. The algorithm for a spherical measurement configuration has recently been reported in another paper. Here, we extend the reconstruction algorithms to planar and cylindrical measurement configurations. First, we generalize the rigorous reconstruction formulas by employing Green's function technique. Then, in order to detect small (compared with the measurement geometry) but deeply buried objects, we can simplify the formulas when two practical conditions exist: 1) that the high-frequency components of the thermoacoustic signals contribute more to the spatial resolution than the low-frequency ones, and 2) that the detecting distances between the thermoacoustic sources and the detecting transducers are much greater than the wavelengths of the high-frequency thermoacoustic signals (i.e., those that are useful for imaging). The simplified formulas are computed with temporal back projections and coherent summations over spherical surfaces using certain spatial weighting factors. We refer to these reconstruction formulas as modified back projections. Numerical results are given to illustrate the validity of these algorithms.

Contrast Mechanisms in Polarization-sensitive Mueller-matrix Optical Coherence Tomography and Application in Burn Imaging

Applied Optics. Sep, 2003  |  Pubmed ID: 12962400

We investigate the various contrast mechanisms provided by polarization-sensitive (PS) Mueller-matrix optical coherence tomography (OCT). Our PS multichannel Mueller-matrix OCT is the first, to our knowledge, to offer simultaneously comprehensive polarization-contrast mechanisms, including the amplitude of birefringence, the orientation of birefringence, and the diattenuation in addition to the polarization-independent intensity contrast, all of which can be extracted from the measured Jones or the equivalent Mueller matrix. Theoretical analysis shows that when diattenuation is negligible, the round-trip Jones matrix represents a linear retarder, which is the foundation of conventional PS-OCT, and can be calculated with a single incident polarization state, although the one-way Jones matrix generally represents an elliptical retarder; otherwise, two incident polarization states are needed. The experimental results obtained from rat skin samples, which conform well with the histology, show that Mueller OCT provides complementary structural and functional information on biological samples and reveal that polarization contrast is more sensitive to thermal degeneration of biological tissue than amplitude-based contrast. Thus, Mueller OCT has significant potential for application in the noninvasive assessment of burn depth.

Three-dimensional Laser-induced Photoacoustic Tomography of Mouse Brain with the Skin and Skull Intact

Optics Letters. Oct, 2003  |  Pubmed ID: 14514085

Three-dimensional laser-induced photoacoustic tomography, also referred to as optoacoustic tomography, is developed to image animal brain structures noninvasively with the skin and skull intact. This imaging modality combines the advantages of optical contrast and ultrasonic resolution. The distribution of optical absorption in a mouse brain is imaged successfully. The intrinsic optical contrast reveals not only blood vessels but also other detailed brain structures, such as the cerebellum, hippocampus, and ventriculi lateralis. The spatial resolution is primarily diffraction limited by the received photoacoustic waves. Imaged structures of the brain at different depths match the corresponding histological pictures well.

Effects of Acoustic Heterogeneity in Breast Thermoacoustic Tomography

IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control. Sep, 2003  |  Pubmed ID: 14561030

The effects of wavefront distortions induced by acoustic heterogeneities in breast thermoacoustic tomography (TAT) are studied. Amplitude distortions are shown to be insignificant for different scales of acoustic heterogeneities. For wavelength-scale, or smaller, heterogeneities, amplitude distortion of the wavefront is minor as a result of diffraction when the detectors are placed in the far field of the heterogeneities. For larger-scale heterogeneities at the parenchyma wall, by using a ray approach (geometric optics), we show that no refraction-induced multipath interference occurs and, consequently, that no severe amplitude distortion, such as is found in ultrasound tomography, exists. Next, we consider the effects of phase distortions (errors in time-of-flight) in our numerical studies. The numerical results on the spreads of point sources and boundaries caused by the phase distortions are in good agreement with the proposed formula. After that, we demonstrate that the blurring of images can be compensated for by using the distribution of acoustic velocity in the tissues in the reconstructions. The effects of the errors in the acoustical velocities on this compensation also are investigated. An approach to implement the compensation using only TAT data is proposed. Lastly, the differences in the effects of acoustic heterogeneity and the generation of speckles in breast TAT and breast ultrasound imaging are discussed.

Polarized Light Propagation Through Scattering Media: Time-resolved Monte Carlo Simulations and Experiments

Journal of Biomedical Optics. Oct, 2003  |  Pubmed ID: 14563198

A study of polarized light transmitted through randomly scattering media of a polystyrene-microsphere solution is described. Temporal profiles of the Stokes vectors and the degree of polarization are measured experimentally and calculated theoretically based on a Monte Carlo technique. The experimental results match the theoretical results well, which demonstrates that the time-resolved Monte Carlo technique is a powerful tool that can contribute to the understanding of polarization propagation in biological tissue. Analysis based on the Stokes-Mueller formalism and the Mie theory shows that the first scattering event determines the major spatial patterns of the transmitted Stokes vectors. When an area detected at the output surface of a turbid medium is circularly symmetrical about the incident beam, the temporal profile of the transmitted light is independent of the incident polarization state. A linear relationship between the average order of the scatters and the light propagation time can be used to explain the exponential decay of the degree of polarization of transmitted light.

Time Reversal and Its Application to Tomography with Diffracting Sources

Physical Review Letters. Jan, 2004  |  Pubmed ID: 14753876

An exact time-domain method is proposed to time reverse a transient scalar wave using only the field measured on an arbitrary closed surface enclosing the initial source. Under certain conditions, a time-reversed field can be approximated by retransmitting the measured signals in a reversed temporal order. Exact reconstruction for three-dimensional broadband diffraction tomography (a linearized inverse scattering problem) is proposed by time-reversing the measured field back to the time when each secondary source is excited. The algorithm is verified by a numerical simulation. Extension to the case using Green's function in a heterogeneous medium is discussed.

Signal Dependence and Noise Source in Ultrasound-modulated Optical Tomography

Applied Optics. Feb, 2004  |  Pubmed ID: 15008535

A Monte Carlo modeling technique was used to simulate ultrasound-modulated optical tomography in inhomogeneous scattering media. The contributions from two different modulation mechanisms were included in the simulation. Results indicate that ultrasound-modulated optical signals are much more sensitive to small embedded objects than unmodulated intensity signals. The differences between embedded absorption and scattering objects in the ultrasound-modulated optical signals were compared. The effects of neighboring inhomogeneity and background optical properties on the ultrasound-modulated optical signals were also studied. We analyzed the signal-to-noise ratio in the experiment and found that the major noise source is the speckle noise caused by small particle movement within the biological tissue sample. We studied this effect by incorporating a Brownian motion factor in the simulation.

Noninvasive Photoacoustic Angiography of Animal Brains in Vivo with Near-infrared Light and an Optical Contrast Agent

Optics Letters. Apr, 2004  |  Pubmed ID: 15072373

Optical contrast agents have been widely applied to enhance the sensitivity and specificity of optical imaging with near-infrared (NIR) light. However, because of the overwhelming scattering of light in biological tissues, the spatial resolution of traditional optical imaging degrades drastically as the imaging depth increases. Here, for the first time to our knowledge, we present noninvasive photoacoustic angiography of animal brains in vivo with NIR light and an optical contrast agent. When indocyanine green polyethylene glycol, a novel absorption dye with prolonged clearance, is injected into the circulatory system of a rat, it obviously enhances the absorption contrast between the blood vessels and the background tissues. Because NIR light can penetrate deep into the brain tissues through the skin and skull, we are able to successfully reconstruct the vascular distribution in the rat brain from the photoacoustic signals. On the basis of differential optical absorption with and without contrast enhancement, a photoacoustic angiograph of a rat brain is acquired that matches the anatomical photograph well and exhibits high spatial resolution and a much-reduced background. This new technology demonstrates the potential for dynamic and molecular biomedical imaging.

Reconstructions in Limited-view Thermoacoustic Tomography

Medical Physics. Apr, 2004  |  Pubmed ID: 15124989

The limited-view problem is studied for thermoacoustic tomography, which is also referred to as photoacoustic or optoacoustic tomography depending on the type of radiation for the induction of acoustic waves. We define a "detection region," within which all points have sufficient detection views. It is explained analytically and shown numerically that the boundaries of any objects inside this region can be recovered stably. Otherwise some sharp details become blurred. One can identify in advance the parts of the boundaries that will be affected if the detection view is insufficient. If the detector scans along a circle in a two-dimensional case, acquiring a sufficient view might require covering more than a pi-, or less than a pi-arc of the trajectory depending on the position of the object. Similar results hold in a three-dimensional case. In order to support our theoretical conclusions, three types of reconstruction methods are utilized: a filtered backprojection (FBP) approximate inversion, which is shown to work well for limited-view data, a local-tomography-type reconstruction that emphasizes sharp details (e.g., the boundaries of inclusions), and an iterative algebraic truncated conjugate gradient algorithm used in conjunction with FBP. Computations are conducted for both numerically simulated and experimental data. The reconstructions confirm our theoretical predictions.

Formulation of Photon Diffusion from Spherical Bioluminescent Sources in an Infinite Homogeneous Medium

Biomedical Engineering Online. May, 2004  |  Pubmed ID: 15125780

The bioluminescent enzyme firefly luciferase (Luc) or variants of green fluorescent protein (GFP) in transformed cells can be effectively used to reveal molecular and cellular features of neoplasia in vivo. Tumor cell growth and regression in response to various therapies can be evaluated by using bioluminescent imaging. In bioluminescent imaging, light propagates in highly scattering tissue, and the diffusion approximation is sufficiently accurate to predict the imaging signal around the biological tissue. The numerical solutions to the diffusion equation take large amounts of computational time, and the studies for its analytic solutions have attracted more attention in biomedical engineering applications.

Multiple-bandwidth Photoacoustic Tomography

Physics in Medicine and Biology. Apr, 2004  |  Pubmed ID: 15128208

Photoacoustic tomography, also referred to as optoacoustic tomography, employs short laser pulses to generate ultrasonic waves in biological tissues. The reconstructed images can be characterized by the convolution of the structure of samples, the laser pulse and the impulse response of the ultrasonic transducer used for detection. Although the laser-induced ultrasonic waves cover a wide spectral range, a single transducer can receive only part of the spectrum because of its limited bandwidth. To systematically analyse this problem, we constructed a photoacoustic tomographic system that uses multiple ultrasonic transducers simultaneously, each at a different central frequency. The photoacoustic images associated with the different transducers were compared and analysed. The system was tested by imaging both mouse brains and phantom samples. The vascular vessels in the brain were revealed by all of the transducers, but the image resolutions differed. The higher frequency detectors provided better image resolution while the lower frequency detectors delineated the major structural traits with a higher signal-noise ratio.

Skin Cancer Detection by Spectroscopic Oblique-incidence Reflectometry: Classification and Physiological Origins

Applied Optics. May, 2004  |  Pubmed ID: 15130003

Data obtained from 102 skin lesions in vivo by spectroscopic oblique-incidence reflectometry were analyzed. The participating physicians initially divided the skin lesions into two visually distinguishable groups based on the lesions' melanocytic conditions. Group 1 consisted of the following two cancerous and benign subgroups: (1) basal cell carcinomas and squamous cell carcinomas and (2) benign actinic keratoses, seborrheic keratoses, and warts. Group 2 consisted of (1) dysplastic nevi and (2) benign common nevi. For each group, a bootstrap-based Bayes classifier was designed to separate the benign from the dysplastic or cancerous tissues. A genetic algorithm was then used to obtain the most effective combination of spatiospectral features for each classifier. The classifiers, tested with prospective blind studies, reached statistical accuracies of 100% and 95% for groups 1 and 2, respectively. Properties that related to cell-nuclear size, to the concentration of oxyhemoglobin, and to the concentration of deoxyhemoglobin as well as the derived concentration of total hemoglobin and oxygen saturation were defined to explain the origins of the classification outcomes.

A Mouse Optical Simulation Environment (MOSE) to Investigate Bioluminescent Phenomena in the Living Mouse with the Monte Carlo Method

Academic Radiology. Sep, 2004  |  Pubmed ID: 15350584

As an important part of bioluminescence tomography, which is a newly developed optical imaging modality, mouse optical simulation environment (MOSE) is developed to simulate bioluminescent phenomena in the living mouse and to predict bioluminescent signals detectable outside the mouse. This simulator is dedicated to small animal optical imaging based on bioluminescence.

Determination of Local Polarization Properties of Biological Samples in the Presence of Diattenuation by Use of Mueller Optical Coherence Tomography

Optics Letters. Oct, 2004  |  Pubmed ID: 15532281

A unique feature of polarization-sensitive Mueller optical coherence tomography is that, by measuring Jones or Mueller matrices, it can reveal the complete polarization properties of biological samples, even in the presence of diattenuation. We map local polarization properties for the first time to our knowledge by using polar decomposition in combination with least-squares fitting to differentiate measured integrated Jones matrices with respect to depth. We also introduce the new concept of dual attenuation coefficients to characterize diattenuation per unit infinitesimal length in tissues. We experimentally verify the algorithm using measurements of a section of porcine tendon and the septum of a rat heart.

Comparison of Human Skin Opto-thermal Response to Near-infrared and Visible Laser Irradiations: a Theoretical Investigation

Physics in Medicine and Biology. Nov, 2004  |  Pubmed ID: 15584524

Near-infrared wavelengths are absorbed less by epidermal melanin, and penetrate deeper into human skin dermis and blood than visible wavelengths. Therefore, laser irradiation using near-infrared wavelengths may improve the therapeutic outcome of cutaneous hyper-vascular malformations in moderately to heavily pigmented skin patients and those with large-sized blood vessels or blood vessels extending deeply into the skin. A mathematical model composed of a Monte Carlo algorithm to estimate the distribution of absorbed light, numerical solution of a bio-heat diffusion equation to calculate the transient temperature distribution, and a damage integral based on an empirical Arrhenius relationship to quantify the tissue damage was utilized to investigate the optothermal response of human skin to near-infrared and visible laser irradiations in conjunction with cryogen spray cooling. In addition, the thermal effects of a single continuous laser pulse and micropulse-composed laser pulse profiles were compared. Simulation results indicated that a 940 nm wavelength induces improved therapeutic outcome compared with a 585 and 595 nm wavelengths for the treatment of patients with large-sized blood vessels and moderately to heavily pigmented skin. On the other hand, a 585 nm wavelength shows the best efficacy in treating small-sized blood vessels, as characterized by the largest laser-induced blood vessel damage depth compared with 595 and 940 nm wavelengths. Dermal blood content has a considerable effect on the threshold incident dosage for epidermal damage, while the effect of blood vessel size is minimal. For the same macropulse duration and incident dosage, a micropulse-composed pulse profile results in higher peak temperature at the basal layer of skin epidermis than an ideal single continuous pulse profile.

High-resolution Ultrasound-modulated Optical Tomography in Biological Tissues

Optics Letters. Dec, 2004  |  Pubmed ID: 15605500

We present a novel implementation of high-resolution ultrasound-modulated optical tomography that, based on optical contrast, can image several millimeters deep into soft biological tissues. A long-cavity confocal Fabry-Perot interferometer, which provides a large etendue and a short response time, was used to detect the ultrasound-modulated coherent light that traversed the scattering biological tissue. Using 15-MHz ultrasound, we imaged with high-contrast light-absorbing structures placed >3 mm below the surface of chicken breast tissue. The resolution along the axial and the lateral directions with respect to the ultrasound propagation direction was better than 70 and 120 microm, respectively. The resolution can be scaled down further by use of higher ultrasound frequencies. This technology is complementary to other imaging technologies, such as confocal microscopy and optical-coherence tomography, and has the potential for broad biomedical applications.

High-resolution Ultrasound-aided Biophotonic Imaging

Conference Proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference. 2004  |  Pubmed ID: 17271539

Optical contrast is sensitive to functional parameters, including the oxygen saturation and total concentration of hemoglobin, in biological tissues. However, due to the overwhelming scattering encountered by light in tissues, traditional optical modalities cannot provide satisfactory spatial resolution beyond the ballistic (a few hundred microns) and quasiballistic (1-2 mm) regimes. Photoacoustic tomography is based on the high optical contrast yet utilizing the high ultrasonic resolution. Our work in this emerging area of research will be summarized in this invited talk. In this technology, a diffraction-based inverse-source problem is solved in the image reconstruction, for which we developed the rigorous reconstruction theory. We implemented a prototype and accomplished noninvasive transdermal and transcranial functional imaging of small-animal brains in vivo. Change in the cerebral blood oxygenation of a rat, as a result of the alternation from hyperoxia to hypoxia, was imaged successfully.

Modulation of Multiply Scattered Coherent Light by Ultrasonic Pulses: an Analytical Model

Physical Review. E, Statistical, Nonlinear, and Soft Matter Physics. Sep, 2005  |  Pubmed ID: 16241605

We present an analytical solution for the acousto-optical modulation of multiply scattered light in a medium irradiated with a train of ultrasound pulses. Previous theory is extended to cases where the ultrasound-induced optical phase increments between the different scattering events are strongly correlated, and it is shown that the approximate similarity relation still holds. The relation between the ultrasound induced motions of the background fluid and the optical scatterers is generalized, and it is shown that correlation exists between the optical phase increments that are due to the scatterer movement and the optical phase increments that are due to the modulation of the optical index of refraction. Finally, it is shown that compared with the spectrum of ultrasound pulses, the power spectral density of acousto-optically modulated light is strongly attenuated at the higher ultrasound frequencies.

Synchronous Self-elimination of Autocorrelation Interference in Fourier-domain Optical Coherence Tomography

Optics Letters. Nov, 2005  |  Pubmed ID: 16279475

We have developed a new algorithm and configuration for self-eliminating the autocorrelation of the object wave in Fourier-domain optical coherence tomography. A self-interferogram of the object wave is acquired synchronously with the standard interferogram of the recombined object and reference waves. The former is then subtracted from the latter after Fourier transformation. The algorithm is validated by numerical simulation and by experimental measurement of a U.S. Air Force target and a feline eye.

Universal Back-projection Algorithm for Photoacoustic Computed Tomography

Physical Review. E, Statistical, Nonlinear, and Soft Matter Physics. Jan, 2005  |  Pubmed ID: 15697763

We report results of a reconstruction algorithm for three-dimensional photoacoustic computed tomography. A universal back-projection formula is presented for three types of imaging geometries: planar, spherical, and cylindrical surfaces. A solid-angle weighting factor is introduced in the back-projection formula to compensate for the variations of detection views. A method for implementing this algorithm is described. Numerical simulation is used to demonstrate the performance of the algorithm.

Half-time Image Reconstruction in Thermoacoustic Tomography

IEEE Transactions on Medical Imaging. Feb, 2005  |  Pubmed ID: 15707246

Thermoacoustic tomography (TAT) is an emerging imaging technique with great potential for a wide range of biomedical imaging applications. In this paper, we propose and investigate reconstruction approaches for TAT that are based on the half-time reflectivity tomography paradigm. We reveal that half-time reconstruction approaches permit for the explicit control of statistically complementary information that can result in the optimal reduction of image variances. We also show that half-time reconstruction approaches can mitigate image artifacts due to heterogeneous acoustic properties of an object. Reconstructed images and numerical results produced from simulated and experimental TAT measurement data are employed to demonstrate these effects.

Imaging of High-intensity Focused Ultrasound-induced Lesions in Soft Biological Tissue Using Thermoacoustic Tomography

Medical Physics. Jan, 2005  |  Pubmed ID: 15719948

An imaging technology, thermoacoustic tomograpy (TAT), was applied to the visualization of high-intensity focused ultrasound (HIFU)-induced lesions. A single, spherically focused ultrasonic transducer, operating at a central frequency of approximately 4 MHz, was used to generate a HIFU field in fresh porcine muscle. Microwave pulses from a 3-GHz microwave generator were then employed to generate thermoacoustic sources in this tissue sample. The thermoacoustic signals were detected by an unfocused ultrasonic transducer that was scanned around the sample. To emphasize the boundaries between the lesion and its surrounding tissue, a local-tomography-type reconstruction method was applied to reconstruct the TAT images of the lesions. Good contrast was obtained between the lesion and the tissue surrounding it. Gross pathologic photographs of the tissue samples confirmed the TAT images. This work demonstrates that TAT may potentially be used to image HIFU-induced lesions in biological tissues.

Imaging of Tumor Angiogenesis in Rat Brains in Vivo by Photoacoustic Tomography

Applied Optics. Feb, 2005  |  Pubmed ID: 15751858

Green laser pulses at a wavelength of 532 nm from a Q-switched Nd:YAG laser were employed as irradiation sources for photoacoustic tomography (PAT). The vascular structure of the brain was imaged clearly, with optimal contrast, because blood has strong absorption near this wavelength. The photoacoustic images of rat brain tumors in this study clearly reveal the angiogenesis that is associated with tumors. Brain tumors can be identified based on the distorted vascular architecture of brain tumorigenesis and related vascular changes, such as hemorrhage. This research demonstrates that PAT can potentially provide a powerful tool for small-animal biological research.

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.

Deeply Penetrating Photoacoustic Tomography in Biological Tissues Enhanced with an Optical Contrast Agent

Optics Letters. Mar, 2005  |  Pubmed ID: 15789718

Photoacoustic tomography (PAT) in a circular scanning configuration was developed to image deeply embedded optical heterogeneity in biological tissues. While the optical penetration was maximized with near-infrared laser pulses of 800-nm wavelength, the optical contrast was enhanced by Indocyanine Green (ICG) dye whose absorption peak matched the laser wavelength. This optimized PAT was able to image objects embedded at depths of as much as 5.2 cm, 6.2 times the 1/e optical penetration depth, in chicken breast muscle at a resolution of <780 microm and a sensitivity of <7 pmol of ICG in blood. The resolution was found to deteriorate slowly with increasing imaging depth. The effects of detection bandwidth on the quality of images acquired simultaneously by four different ultrasonic transducers are described.

In Vivo Dark-field Reflection-mode Photoacoustic Microscopy

Optics Letters. Mar, 2005  |  Pubmed ID: 15791997

Reflection-mode photoacoustic microscopy with dark-field laser pulse illumination and high-numerical-aperture ultrasonic detection is designed and implemented in noninvasively imaged blood vessels in the skin in vivo. Dark-field optical illumination minimizes the interference caused by strong photoacoustic signals from superficial structures. A high-numerical-aperture acoustic lens provides high lateral resolution, 45-120 microm in this system. A broadband ultrasonic detection system provides high axial resolution, estimated to be approximately 15 microm. The optical illumination and ultrasonic detection are in a coaxial confocal configuration for optimal image quality. The system is capable of imaging optical-absorption contrast as deep as 3 mm in biological tissue.

Weighted Expectation Maximization Reconstruction Algorithms for Thermoacoustic Tomography

IEEE Transactions on Medical Imaging. Jun, 2005  |  Pubmed ID: 15957603

Thermoacoustic tomography (TAT) is an emerging imaging technique with potential for a wide range of biomedical imaging applications. In this correspondence, we propose an infinite family of weighted expectation maximization (EM) algorithms for reconstruction of images from temporally truncated TAT measurement data. The weighted EM algorithms are equivalent mathematically to the conventional EM algorithm, but are shown to propagate data inconsistencies in different ways. Using simulated and experimental TAT measurement data, we demonstrate that suitable choices of weighted EM algorithms can effectively mitigate image artifacts that are attributable to temporal truncation of the TAT data function.

Fiber-based Polarization-sensitive Mueller Matrix Optical Coherence Tomography with Continuous Source Polarization Modulation

Applied Optics. Sep, 2005  |  Pubmed ID: 16161660

We report on a new configuration of fiber-based polarization-sensitive Mueller matrix optical coherence tomography that permits the acquisition of the round-trip Jones matrix of a biological sample using only one light source and a single depth scan. In this new configuration, a polarization modulator is used in the source arm to continuously modulate the incident polarization state for both the reference and the sample arms. The Jones matrix of the sample can be calculated from the two frequency terms in the two detection channels. The first term is modulated by the carrier frequency, which is determined by the longitudinal scanning mechanism, whereas the other term is modulated by the beat frequency between the carrier frequency and the second harmonic of the modulation frequency of the polarization modulator. One important feature of this system is that, for the first time to our knowledge, the Jones matrix of the sample can be calculated with a single detection channel and a single measurement when diattenuation is negligible. The system was successfully tested by imaging both standard polarization elements and biological samples.

Thermoacoustic and Photoacoustic Tomography of Thick Biological Tissues Toward Breast Imaging

Technology in Cancer Research & Treatment. Oct, 2005  |  Pubmed ID: 16173826

Microwave-based thermoacoustic tomography (TAT) and laser-based photoacoustic tomography (PAT) in a circular scanning configuration were both developed to image deeply seated lesions and objects in biological tissues. Because malignant breast tissue absorbs microwaves more strongly than benign breast tissue, cancers were imaged with good spatial resolution and contrast by TAT in human breast mastectomy specimens. Based on the intrinsic optical contrast between blood and chicken breast muscle, an embedded blood object that was 5 cm deep in the tissue was also detected using PAT at a wavelength of 1064 nm.

Improved in Vivo Photoacoustic Microscopy Based on a Virtual-detector Concept

Optics Letters. Feb, 2006  |  Pubmed ID: 16496891

Recently an in vivo high-resolution backward-mode photoacoustic microscope was developed that shows potential for applications in dermatology and related cancer research. However, the limited depth of focus of the large-numerical-aperture (NA) ultrasonic lens employed in this system causes the image quality to deteriorate significantly in the out-of-focus region. To solve this problem, we devised and explored, for the first time to our knowledge, a virtual-detector-based synthetic-aperture focusing technique, combined with coherence weighting, for photoacoustic microscopy with such a large-NA transducer. Images of phantoms show that the proposed technique improves the -6 dB lateral resolution from 49-379 to 46-53 microm and increases the signal-to-noise ratio by up to 29 dB, depending on the distance from the ultrasonic focal point. In vivo experiments show that the technique also provides a clearer representation of the vascular distribution in the rat's scalp.

Rhesus Monkey Brain Imaging Through Intact Skull with Thermoacoustic Tomography

IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control. Mar, 2006  |  Pubmed ID: 16555762

Two-dimensional microwave-induced thermoacoustic tomography (TAT) is applied to imaging the Rhesus monkey brain through the intact skull. To reduce the wavefront distortion caused by the skull, only the low-frequency components of the thermoacoustic signals (< 1 MHz) are used to reconstruct the TAT images. The methods of signal processing and image reconstruction are validated by imaging a lamb kidney. The resolution of the system is found to be 4 mm when we image a 1-month-old monkey head containing inserted needles. We also image the coronal and axial sections of a 7-month-old monkey head. Brain features that are 3 cm deep in the head are imaged clearly. Our results demonstrate that TAT has potential for use in portable, cost-effective imagers for pediatric brains.

Noninvasive Imaging of Hemoglobin Concentration and Oxygenation in the Rat Brain Using High-resolution Photoacoustic Tomography

Journal of Biomedical Optics. Mar-Apr, 2006  |  Pubmed ID: 16674205

Simultaneous transcranial imaging of two functional parameters, the total concentration of hemoglobin and the hemoglobin oxygen saturation, in the rat brain in vivo is realized noninvasively using laser-based photoacoustic tomography (PAT). As in optical diffusion spectroscopy, PAT can assess the optical absorption of endogenous chromophores, e.g., oxygenated and deoxygenated hemoglobins, at multiple optical wavelengths. However, PAT can provide high spatial resolution because its resolution is diffraction-limited by photoacoustic signals rather than by optical diffusion. Laser pulses at two wavelengths are used sequentially to acquire photoacoustic images of the vasculature in the cerebral cortex of a rat brain through the intact skin and skull. The distributions of blood volume and blood oxygenation in the cerebral cortical venous vessels, altered by systemic physiological modulations including hyperoxia, normoxia, and hypoxia, are visualized successfully with satisfactory spatial resolution. This technique, with its prominent sensitivity to endogenous contrast, can potentially contribute to the understanding of the interrelationship between neural, hemodynamic, and metabolic activities in the brain.

Correlation Transfer and Diffusion of Ultrasound-modulated Multiply Scattered Light

Physical Review Letters. Apr, 2006  |  Pubmed ID: 16712231

We develop a temporal correlation transfer equation (CTE) and a temporal correlation diffusion equation (CDE) for ultrasound-modulated multiply scattered light. These equations can be applied to an optically scattering medium with embedded optically scattering and absorbing objects to calculate the power spectrum of light modulated by a nonuniform ultrasound field. We present an analytical solution based on the CDE and Monte Carlo simulation results for light modulated by a cylinder of ultrasound in an optically scattering slab. We further validate with experimental measurements the numerical calculations for an actual ultrasound field. The CTE and CDE are valid for moderate ultrasound pressures and on a length scale comparable with the optical transport mean-free path. These equations should be applicable to a wide spectrum of conditions for ultrasound-modulated optical tomography of soft biological tissues.

Temperature Distribution in Selective Laser-tissue Interaction

Journal of Biomedical Optics. May-Jun, 2006  |  Pubmed ID: 16822080

Selective photothermal interaction using dye enhancement has proven to be effective in minimizing surrounding tissue damage and delivering energy to target tissue. During laser irradiation, the process of photon absorption and thermal energy diffusion in the target tissue and its surrounding tissue are crucial. Such information allows the selection of proper operating parameters such as dye concentrations, laser power, and exposure time for optimal therapeutic effect. Combining the Monte Carlo method for energy absorption and the finite difference method for heat diffusion, the temperature distributions in target tissue and surrounding tissue in dye enhanced laser photothermal interaction are obtained. Different tissue configurations and dye enhancement are used in the simulation, and different incident beam sizes are also used to determine optimum beam sizes for various tissue configurations. Our results show that the algorithm developed in this study could predict the thermal outcome of laser irradiation. Our simulation indicates that with appropriate absorption enhancement of the target tissue, the temperature in the target tissue and in the surrounding tissue can be effectively controlled. This method can be used for optimization of lesion treatment using laser photothermal interactions. It may also provide guidance for laser immunotherapy in cancer treatment, since the immunological responses are believed to be related to tissue temperature changes.

Three-dimensional Imaging of Skin Melanoma in Vivo by Dual-wavelength Photoacoustic Microscopy

Journal of Biomedical Optics. May-Jun, 2006  |  Pubmed ID: 16822081

Dual-wavelength reflection-mode photoacoustic microscopy is used to noninvasively obtain three-dimensional (3-D) images of subcutaneous melanomas and their surrounding vasculature in nude mice in vivo. The absorption coefficients of blood and melanin-pigmented melanomas vary greatly relative to each other at these two optical wavelengths (764 and 584 nm). Using high-resolution and high-contrast photoacoustic imaging in vivo with a near-infrared (764-nm) light source, the 3-D melanin distribution inside the skin is imaged, and the maximum thickness of the melanoma (approximately 0.5 mm) is measured. The vascular system surrounding the melanoma is also imaged with visible light (584 nm) and the tumor-feeding vessels found. This technique can potentially be used for melanoma diagnosis, prognosis, and treatment planning.

Functional Photoacoustic Microscopy for High-resolution and Noninvasive in Vivo Imaging

Nature Biotechnology. Jul, 2006  |  Pubmed ID: 16823374

Although optical absorption is strongly associated with the physiological status of biological tissue, existing high-resolution optical imaging modalities, including confocal microscopy, two-photon microscopy and optical coherence tomography, do not sense optical absorption directly. Furthermore, optical scattering prevents these methods from imaging deeper than approximately 1 mm below the tissue surface. Here we report functional photoacoustic microscopy (fPAM), which provides multiwavelength imaging of optical absorption and permits high spatial resolution beyond this depth limit with a ratio of maximum imaging depth to depth resolution greater than 100. Reflection mode, rather than orthogonal or transmission mode, is adopted because it is applicable to more anatomical sites than the others. fPAM is demonstrated with in vivo imaging of angiogenesis, melanoma, hemoglobin oxygen saturation (sO2) of single vessels in animals and total hemoglobin concentration in humans.

A Born-type Approximation Method for Bioluminescence Tomography

Medical Physics. Mar, 2006  |  Pubmed ID: 16878571

In this paper, we present a Born-type approximation method for bioluminescence tomography (BLT), which is to reconstruct an internal bioluminescent source from the measured bioluminescent signal on the external surface of a small animal. Based on the diffusion approximation for the photon propagation in biological tissue, this BLT method utilizes the Green function to establish a linear relationship between the measured bioluminescent signal and the internal bioluminescent source distribution. The Green function can be modified to describe a heterogeneous medium with an arbitrary boundary using the Born approximation. The BLT reconstruction is formulated in a linear least-squares optimization framework with simple bounds constraint. The performance of this method is evaluated in numerical simulation and phantom experiments.

Intense Acoustic Bursts As a Signal-enhancement Mechanism in Ultrasound-modulated Optical Tomography

Optics Letters. Aug, 2006  |  Pubmed ID: 16880843

Biophotonic imaging with ultrasound-modulated optical tomography (UOT) promises ultrasonically resolved imaging in biological tissues. A key challenge in this imaging technique is a low signal-to-noise ratio (SNR). We show significant UOT signal enhancement by using intense time-gated acoustic bursts. A CCD camera captured the speckle pattern from a laser-illuminated tissue phantom. Differences in speckle contrast were observed when ultrasonic bursts were applied, compared with when no ultrasound was applied. When CCD triggering was synchronized with burst initiation, acoustic-radiation-force-induced displacements were detected. To avoid mechanical contrast in UOT images, the CCD camera acquisition was delayed several milliseconds until transient effects of acoustic radiation force attenuated to a satisfactory level. The SNR of our system was sufficiently high to provide an image pixel per acoustic burst without signal averaging. Because of the substantially improved SNR, the use of intense acoustic bursts is a promising signal enhancement strategy for UOT.

In Vivo Three-dimensional Photoacoustic Tomography of a Whole Mouse Head

Optics Letters. Aug, 2006  |  Pubmed ID: 16880853

An in vivo photoacoustic imaging system was designed and implemented to image the entire small animal head. A special scanning gantry was designed to enable in vivo imaging in coronal cross sections with high contrast and good spatial resolution for the first time to our knowledge. By use of a 2.25 MHz ultrasonic transducer with a 6 mm diameter active element, an in-plane radial resolution of approximately 312 microm was achieved. Deeply seated arterial and venous vessels in the head measuring up to 1.7 cm in diameter were simultaneously imaged in vivo with 804 nm wavelength laser excitation of photoacoustic waves.

Stochastic Explanation of Speckle Contrast Detection in Ultrasound-modulated Optical Tomography

Physical Review. E, Statistical, Nonlinear, and Soft Matter Physics. Jun, 2006  |  Pubmed ID: 16906877

Ultrasound-modulated optical tomography is an imaging technique that detects ultrasonically tagged light in optically turbid media to obtain images with optical contrast and ultrasonic spatial resolution. A CCD-camera-based speckle contrast detection scheme has been introduced previously to detect modulated light emerging from the ultrasonic sample volume. Differences in speckle contrast were experimentally observed when ultrasound was applied compared to when it was not. In this paper we provide an analytic explanation for this phenomenon and connect speckle statistics with ultrasonic field parameters. The theory predicts that speckle contrast changes linearly with applied acoustic intensity. This prediction is experimentally validated for both 1 and ultrasound. Signal dependence on ultrasound frequency is discussed.

Correlation Transfer Equation for Ultrasound-modulated Multiply Scattered Light

Physical Review. E, Statistical, Nonlinear, and Soft Matter Physics. Sep, 2006  |  Pubmed ID: 17025775

In this paper, we develop a temporal correlation transfer equation (CTE) for ultrasound-modulated multiply scattered light. The equation can be used to obtain the time-varying specific intensity of light produced by a nonuniform ultrasound field in optically scattering media that have a heterogeneous distribution of optical parameters. We also develop a Monte Carlo algorithm that can provide the spatial distribution of the optical power spectrum in optically scattering media with focused ultrasound fields, and heterogeneous distributions of optically scattering and absorbing objects. Derivation of the CTE is based on the ladder diagram approximation of the Bethe-Salpeter equation that assumes moderate ultrasound pressures. We expect the CTE to be applicable to a wide spectrum of conditions in the ultrasound-modulated optical tomography of soft biological tissues.

Imaging Acute Thermal Burns by Photoacoustic Microscopy

Journal of Biomedical Optics. Sep-Oct, 2006  |  Pubmed ID: 17092182

The clinical significance of a burn depends on the percentage of total body involved and the depth of the burn. Hence a noninvasive method that is able to evaluate burn depth would be of great help in clinical evaluation. To this end, photoacoustic microscopy is used to determine the depth of acute thermal burns by imaging the total hemoglobin concentration in the blood that accumulates along the boundaries of injuries as a result of thermal damage to the vasculature. We induce acute thermal burns in vivo on pig skin with cautery. Photoacoustic images of the burns are acquired after skin excision. In a burn treated at 175 degrees C for 20 s, the maximum imaged burn depth is 1.73+/-0.07 mm. In burns treated at 150 degrees C for 5, 10, 20, and 30 s, respectively, the trend of increasing maximum burn depth with longer thermal exposure is demonstrated.

In Vivo Volumetric Imaging of Subcutaneous Microvasculature by Photoacoustic Microscopy

Optics Express. Oct, 2006  |  Pubmed ID: 19529315

Photoacoustic microscopy was developed to achieve volumetric imaging of the anatomy and functions of the subcutaneous microvasculature in both small animals and humans in vivo with high spatial resolution and high signal-to-background ratio. By following the skin contour in raster scanning, the ultrasonic transducer maintains focusing in the region of interest. Furthermore, off-focus lateral resolution is improved by using a synthetic-aperture focusing technique based on the virtual point detector concept. Structural images are acquired in both rats and humans, whereas functional images representing hemoglobin oxygen saturation are acquired in rats. After multiscale vesselness filtering, arterioles and venules in the image are separated based on the imaged oxygen saturation levels. Detailed structural information, such as vessel depth and spatial orientation, are revealed by volume rendering.

Thermoacoustic Tomography with Correction for Acoustic Speed Variations

Physics in Medicine and Biology. Dec, 2006  |  Pubmed ID: 17148827

Thermoacoustic tomography (TAT) is a technique that measures microwave-induced thermoacoustic waves at the boundary of biological tissue and generates images of internal microwave absorption distributions from the measurements. Existing reconstruction algorithms for TAT are based on the assumption that the acoustic properties in the tissue are homogeneous. Biological tissue, however, has heterogeneous acoustic properties, which lead to distortion and blurring of small buried objects in the reconstructed images. In this paper we develop a correction method based on ultrasonic transmission tomography (UTT) to improve the image quality of TAT. Numerical simulations and phantom experiments verify the effectiveness of this correction method.

In Vivo Imaging and Characterization of Hypoxia-induced Neovascularization and Tumor Invasion

International Journal of Oncology. Jan, 2007  |  Pubmed ID: 17143511

Hypoxia is a critical event in tumor progression and angiogenesis. Hypoxia can be detected noninvasively by a novel spectroscopic photoacoustic tomography technology (SPAT) and this finding is supported by our molecular biology investigation aimed to elucidate the etiopathogenesis of SPAT detected hypoxia and angiogenesis. The present study provides an integrated approach to define oxygen status (hypoxia) of intracranial tumor xenografts using spectroscopic photoacoustic tomography. Brain tumors can be identified based on their distorted vascular architecture and oxygen saturation (SO2) images. Noninvasive in vivo tumor oxygenation imaging using SPAT is based on the spectroscopic absorption differences between oxyhemoglobin (O2Hb) and deoxyhemoblobin (HHb). Sprague-Dawley rats inoculated intracranially with ENU1564, a carcinogen-induced rat mammary adenocarcinoma cell line, were imaged with SPAT three weeks post inoculation. Proteins important for tumor angiogenesis and invasion were detected in hypoxic brain foci identified by SPAT and were elevated compared with control brain. Immunohistochemistry, Western blotting, and semi-quantitative RT-PCR showed that HIF-1 alpha, VEGF-A, and VEGFR2 (Flk-1) protein and mRNA expression levels were significantly higher (P < 0.05) in brain tumor tissues compared to normal brain. Gelatin zymography and RT-PCR demonstrated the upregulation of MMP-9 in tumor foci compared with brain control. Together these results suggest the critical role of hypoxia in driving tumor angiogenesis and invasion through upregulation of target genes important for these functions. Moreover this report validates our hypothesis that a novel noninvasive technology (SPAT) developed in our laboratory is suitable for detection of tumors, hypoxia, and angiogenesis.

Limitations of Quantitative Photoacoustic Measurements of Blood Oxygenation in Small Vessels

Physics in Medicine and Biology. Mar, 2007  |  Pubmed ID: 17301459

We investigate the feasibility of obtaining accurate quantitative information, such as local blood oxygenation level (sO2), with a spatial resolution of about 50 microm from spectral photoacoustic (PA) measurements. The optical wavelength dependence of the peak values of the PA signals is utilized to obtain the local blood oxygenation level. In our in vitro experimental models, the PA signal amplitude is found to be linearly proportional to the blood optical absorption coefficient when using ultrasonic transducers with central frequencies high enough such that the ultrasonic wavelengths are shorter than the light penetration depth into the blood vessels. For an optical wavelength in the 578-596 nm region, with a transducer central frequency that is above 25 MHz, the sensitivity and accuracy of sO2 inversion is shown to be better than 4%. The effect of the transducer focal position on the accuracy of quantifying blood oxygenation is found to be negligible. In vivo oxygenation measurements of rat skin microvasculature yield results consistent with those from in vitro studies, although factors specific to in vivo measurements, such as the spectral dependence of tissue optical attenuation, dramatically affect the accuracy of sO2 quantification in vivo.

Photorefractive Detection of Tissue Optical and Mechanical Properties by Ultrasound Modulated Optical Tomography

Optics Letters. Mar, 2007  |  Pubmed ID: 17308592

Ultrasound-modulated optical tomography is a developing hybrid imaging modality that combines high optical contrast and good ultrasonic resolution for imaging soft biological tissue. We developed a photorefractive-crystal-based, time-resolved detection scheme with the use of a millisecond long ultrasound burst to image both the optical and the mechanical properties of biological tissues, with improved detection efficiency of ultrasound-tagged photons.

Photoacoustic Imaging of the Microvasculature with a High-frequency Ultrasound Array Transducer

Journal of Biomedical Optics. Jan-Feb, 2007  |  Pubmed ID: 17343475

Visualization of microvascular networks could provide new information about function and disease. We demonstrate the capabilities of a 30-MHz ultrasound array system for photoacoustic microscopy of small (< or = 300 microm) vessels in a rat. 3D images obtained by translating the array in the elevation direction are compared with photographs of excised skin. The system is shown to have 100-microm lateral resolution, 25-microm axial resolution, and 3-mm imaging depth. To our knowledge this is the first report on photoacoustic microscopy of the microvasculature with a high-frequency array transducer. It is anticipated that the system can be used for studying and diagnosing a number of diseases including cancer, atherosclerosis, dermatological disorders, and peripheral microvascular complications in diabetes.

Boundary Conditions in Photoacoustic Tomography and Image Reconstruction

Journal of Biomedical Optics. Jan-Feb, 2007  |  Pubmed ID: 17343502

Recently, the field of photoacoustic tomography has experienced considerable growth. Although several commercially available pure optical imaging modalities, including confocal microscopy, two-photon microscopy, and optical coherence tomography, have been highly successful, none of these technologies can penetrate beyond approximately 1 mm into scattering biological tissues because all of them are based on ballistic and quasiballistic photons. Consequently, heretofore there has been a void in high-resolution optical imaging beyond this depth limit. Photoacoustic tomography has filled this void by combining high ultrasonic resolution and strong optical contrast in a single modality. However, it has been assumed in reconstruction of photoacoustic tomography until now that ultrasound propagates in a boundary-free infinite medium. We present the boundary conditions that must be considered in certain imaging configurations; the associated inverse solutions for image reconstruction are provided and validated by numerical simulation and experiment. Partial planar, cylindrical, and spherical detection configurations with a planar boundary are covered, where the boundary can be either hard or soft. Analogously to the method of images of sources, which is commonly used in forward problems, the ultrasonic detectors are imaged about the boundary to satisfy the boundary condition in the inverse problem.

Ultrasound-modulated Optical Tomography with Intense Acoustic Bursts

Applied Optics. Apr, 2007  |  Pubmed ID: 17356603

Ultrasound-modulated optical tomography (UOT) detects ultrasonically modulated light to spatially localize multiply scattered photons in turbid media with the ultimate goal of imaging the optical properties in living subjects. A principal challenge of the technique is weak modulated signal strength. We discuss ways to push the limits of signal enhancement with intense acoustic bursts while conforming to optical and ultrasonic safety standards. A CCD-based speckle-contrast detection scheme is used to detect acoustically modulated light by measuring changes in speckle statistics between ultrasound-on and ultrasound-off states. The CCD image capture is synchronized with the ultrasound burst pulse sequence. Transient acoustic radiation force, a consequence of bursts, is seen to produce slight signal enhancement over pure ultrasonic-modulation mechanisms for bursts and CCD exposure times of the order of milliseconds. However, acoustic radiation-force-induced shear waves are launched away from the acoustic sample volume, which degrade UOT spatial resolution. By time gating the CCD camera to capture modulated light before radiation force has an opportunity to accumulate significant tissue displacement, we reduce the effects of shear-wave image degradation, while enabling very high signal-to-noise ratios. Additionally, we maintain high-resolution images representative of optical and not mechanical contrast. Signal-to-noise levels are sufficiently high so as to enable acquisition of 2D images of phantoms with one acoustic burst per pixel.

In Vivo Imaging of Subcutaneous Structures Using Functional Photoacoustic Microscopy

Nature Protocols. 2007  |  Pubmed ID: 17446879

Functional photoacoustic microscopy (fPAM) is a hybrid technology that permits noninvasive imaging of the optical absorption contrast in subcutaneous biological tissues. fPAM uses a focused ultrasonic transducer to detect high-frequency photoacoustic (PA) signals. Volumetric images of biological tissues can be formed by two-dimensional raster scanning, and functional parameters can be further extracted from spectral measurements. fPAM is safe and applicable to animals as well as humans. This protocol provides guidelines for parameter selection, system alignment, imaging operation, laser safety and data processing for in vivo fPAM. It currently takes approximately 100 min to carry out this protocol, including approximately 50 min for data acquisition using a 10-Hz pulse-repetition-rate laser system. The data acquisition time, however, can be significantly reduced by using a laser system with a higher pulse repetition rate.

Photoacoustic Imaging of LacZ Gene Expression in Vivo

Journal of Biomedical Optics. Mar-Apr, 2007  |  Pubmed ID: 17477703

In the postgenomic era, imaging techniques are playing an important role in visualizing gene expression in vivo. This work represents the first demonstration of photoacoustic tomography (PAT) for reporter gene imaging. Rats inoculated with 9L/lacZ gliosarcoma tumor cells are imaged with PAT before and after injection of X-gal, a colorimetric assay for the lacZ-encoded enzyme beta-galactosidase. Using far-red optical illumination, the genetically tagged tumors in rats are clearly visualized by PAT following the assay. The spatial resolution is quantified to be less than 400 microm, while 500-nM-level sensitivity is demonstrated. With the future development of new absorption-based reporter gene systems, it is anticipated that photoacoustic technology will provide a valuable tool for molecular imaging research.

Multiple-source Optical Diffusion Approximation for a Multilayer Scattering Medium

Applied Optics. Aug, 2007  |  Pubmed ID: 17694156

A method for improving the accuracy of the optical diffusion theory for a multilayer scattering medium is presented. An infinitesimally narrow incident light beam is replaced by multiple isotropic point sources of different strengths that are placed in the scattering medium along the incident beam. The multiple sources are then used to develop a multilayer diffusion theory. Diffuse reflectance is then computed using the multilayer diffusion theory and compared with accurate data computed by the Monte Carlo method. This multisource method is found to be significantly more accurate than the previous single-source method.

Multi-optical-wavelength Ultrasound-modulated Optical Tomography: a Phantom Study

Optics Letters. Aug, 2007  |  Pubmed ID: 17700760

We used multiple optical wavelengths to study ultrasound-modulated optical tomography (UOT) in tissue phantoms. By using intense acoustic bursts and a CCD camera-based speckle contrast detection technique, we observed variations of the ultrasound-modulated signal at various optical absorptions. The experimental variations were found to be highly correlated with predictions from Monte Carlo simulations. By irradiating the sample at two optical wavelengths, we quantitatively estimated the total concentration and the concentration ratio of double dyes in objects embedded in tissue phantoms. The results suggest that UOT can potentially provide noninvasive functional imaging of the total concentration and oxygen saturation of hemoglobin in biological tissue.

Imaging Optically Scattering Objects with Ultrasound-modulated Optical Tomography

Optics Letters. Aug, 2007  |  Pubmed ID: 17700782

We show the feasibility of imaging objects having different optical scattering coefficients relative to the surrounding scattering medium using ultrasound-modulated optical tomography (UOT). While the spatial resolution depends on ultrasound parameters, the image contrast depends on the difference in scattering coefficient between the object and the surrounding medium. Experimental measurements obtained with a CCD-based speckle contrast detection scheme are in agreement with Monte Carlo simulations and analytical calculations. This study complements previous UOT experiments that demonstrated optical absorption contrast.

Correlation Transfer Equation for Multiply Scattered Light Modulated by an Ultrasonic Pulse

Journal of the Optical Society of America. A, Optics, Image Science, and Vision. Sep, 2007  |  Pubmed ID: 17767248

We develop a temporal correlation transfer equation (CTE) and a Monte Carlo algorithm (MC) for multiply scattered light modulated by an ultrasonic pulse propagating in an optically scattering medium, where the ultrasound field can be nonuniform and the medium can have spatially heterogeneous distribution of optical parameters. The CTE and MC can be used to obtain the time-varying specific intensity and the spatial distribution of the time-dependent power spectral density, respectively, of ultrasound-modulated light. We expect the CTE and MC to be applicable for estimation of contrast and resolution in a wide spectrum of conditions in ultrasound-modulated optical tomography of soft biological tissues.

Photoacoustic Doppler Effect from Flowing Small Light-absorbing Particles

Physical Review Letters. Nov, 2007  |  Pubmed ID: 17995411

From the flow of a suspension of micrometer-scale carbon particles, the photoacoustic Doppler shift is observed. As predicted theoretically, the observed Doppler shift equals half of that in Doppler ultrasound and does not depend on the direction of laser illumination. This new physical phenomenon provides a basis for developing photoacoustic Doppler flowmetry, which can potentially be used for detecting fluid flow in optically scattering media and especially low-speed blood flow of relatively deep microcirculation in biological tissue.

Photoacoustic Tomography of a Rat Cerebral Cortex in Vivo with Au Nanocages As an Optical Contrast Agent

Nano Letters. Dec, 2007  |  Pubmed ID: 18020475

Poly(ethylene glycol)-coated Au nanocages have been evaluated as a potential near-infrared (NIR) contrast agent for photoacoustic tomography (PAT). Previously, Au nanoshells were found to be an effective NIR contrast agent for PAT; however, Au nanocages with their more compact sizes (<50 nm compared to >100 nm for Au nanoshells) and larger optical absorption cross sections should be better suited for in vivo applications. We sequentially injected Au nanocages into the circulatory system of a rat in three administrations and in vivo PAT was conducted immediately prior to the first injection and continued until 5 h after the final injection. A gradual enhancement of the optical absorption in the cerebral cortex, by up to 81%, was observed over the course of the experiment.

Evaluation of the Magneto-optical Effect in Biological Tissue Models Using Optical Coherence Tomography

Journal of Biomedical Optics. Nov-Dec, 2007  |  Pubmed ID: 18163797

For the first time to our knowledge, an experimental evaluation of the Faraday effect-induced polarization rotation in a biological tissue phantom is reported. The rotation of the polarization plane produced in the optical beam propagating through an Intralipid solution was evaluated using polarization-sensitive optical coherence tomography (PS-OCT), and the experimental results closely matched the theoretical values. The angle of rotation is proportional to the traversed path length along the magnetic field and can potentially be used to estimate the actual penetration depth.

Deep Reflection-mode Photoacoustic Imaging of Biological Tissue

Journal of Biomedical Optics. Nov-Dec, 2007  |  Pubmed ID: 18163798

A reflection-mode photoacoustic (PA) imaging system was designed and built to image deep structures in biological tissues. We chose near-infrared laser pulses of 804-nm wavelength for PA excitation to achieve deep penetration. To minimize unwanted surface signals, we adopted dark-field ring-shaped illumination. This imaging system employing a 5-MHz spherically focused ultrasonic transducer provides penetration up to 38 mm in chicken breast tissue. At the 19-mm depth, the axial resolution is 144 microm and the transverse resolution is 560 microm. Internal organs of small animals were imaged clearly.

Photoacoustic Tomography of a Rat Cerebral Cortex with a Ring-based Ultrasonic Virtual Point Detector

Journal of Biomedical Optics. Nov-Dec, 2007  |  Pubmed ID: 18163802

We image a rat cerebral cortex in situ by using a ring-based ultrasonic virtual point detector developed previously. Compared to the image generated by a finite-aperture detector, the image generated by the virtual point detector has a uniformly distributed resolution throughout the imaged area, owing to the lack of aperture effect of the ultrasonic detector. At the periphery of the image, the signal-to-noise ratio of the image obtained by the virtual point detector is also better than that of a finite-aperture detector. Furthermore, the virtual point detector can be scanned inside the brain to improve the local signal-to-noise ratio.

Prospects of Photoacoustic Tomography

Medical Physics. Dec, 2008  |  Pubmed ID: 19175133

Commercially available high-resolution three-dimensional optical imaging modalities-including confocal microscopy, two-photon microscopy, and optical coherence tomography-have fundamentally impacted biomedicine. Unfortunately, such tools cannot penetrate biological tissue deeper than the optical transport mean free path (approximately 1 mm in the skin). Photoacoustic tomography, which combines strong optical contrast and high ultrasonic resolution in a single modality, has broken through this fundamental depth limitation and achieved superdepth high-resolution optical imaging. In parallel, radio frequency-or microwave-induced thermoacoustic tomography is being actively developed to combine radio frequency or microwave contrast with ultrasonic resolution. In this Vision 20/20 article, the prospects of photoacoustic tomography are envisaged in the following aspects: (1) photoacoustic microscopy of optical absorption emerging as a mainstream technology, (2) melanoma detection using photoacoustic microscopy, (3) photoacoustic endoscopy, (4) simultaneous functional and molecular photoacoustic tomography, (5) photoacoustic tomography of gene expression, (6) Doppler photoacoustic tomography for flow measurement, (7) photoacoustic tomography of metabolic rate of oxygen, (8) photoacoustic mapping of sentinel lymph nodes, (9) multiscale photoacoustic imaging in vivo with common signal origins, (10) simultaneous photoacoustic and thermoacoustic tomography of the breast, (11) photoacoustic and thermoacoustic tomography of the brain, and (12) low-background thermoacoustic molecular imaging.

Optical-resolution Photoacoustic Microscopy for in Vivo Imaging of Single Capillaries

Optics Letters. May, 2008  |  Pubmed ID: 18451942

Capillaries, the smallest blood vessels, are the distal end of the vasculature where oxygen and nutrients are exchanged between blood and tissue. Hence, noninvasive imaging of capillaries and function in vivo has long been desired as a window to studying fundamental physiology, such as neurovascular coupling. Existing imaging modalities cannot provide the required sensitivity and spatial resolution. We present in vivo imaging of the microvasculature including single capillaries in mice using optical-resolution photoacoustic microscopy (OR-PAM) developed in our laboratory. OR-PAM provides a lateral resolution of 5 microm and an imaging depth >0.7 mm.

Sentinel Lymph Node Detection Ex Vivo Using Ultrasound-modulated Optical Tomography

Journal of Biomedical Optics. Mar-Apr, 2008  |  Pubmed ID: 18465949

We apply ultrasound-modulated optical tomography (UOT) to image ex-vivo methylene-blue-dyed sentinel lymph nodes embedded in 3.2-cm-thick chicken breast tissues. The UOT system is implemented for the first time using ring-shaped light illumination, intense acoustic bursts, and charge-coupled device (CCD) camera-based speckle contrast detection. Since the system is noninvasive, nonionizing, portable, relatively cost effective, and easy to combine with photoacoustic imaging and single element ultrasonic pulse-echo imaging, UOT can potentially be a good imaging modality for the detection of sentinel lymph nodes in breast cancer staging in vivo.

Photoacoustic Imaging of Biological Tissue with Intensity-modulated Continuous-wave Laser

Journal of Biomedical Optics. Mar-Apr, 2008  |  Pubmed ID: 18465969

We build a photoacoustic imaging system using an intensity-modulated continuous-wave laser source, which is an inexpensive, compact, and durable 120-mW laser diode. The goal is to significantly reduce the costs and sizes of photoacoustic imaging systems. By using a bowl-shaped piezoelectric transducer, whose numerical aperture is 0.85 and resonance frequency is 2.45 MHz, we image biological tissues with a lateral resolution of 0.45 mm, an axial resolution of 1 mm, and an SNR as high as 43 dB.

Curved Array Photoacoustic Tomographic System for Small Animal Imaging

Journal of Biomedical Optics. Mar-Apr, 2008  |  Pubmed ID: 18465970

We present systematic characterization of a photoacoustic imaging system optimized for rapid, high-resolution tomographic imaging of small animals. The system is based on a 128-element ultrasonic transducer array with a 5-MHz center frequency and 80% bandwidth shaped to a quarter circle of 25 mm radius. A 16-channel data-acquisition module and dedicated channel detection electronics enable capture of a 90-deg field-of-view image in less than 1 s and a complete 360-deg scan using sample rotation within 15 s. Measurements on cylindrical phantom targets demonstrate a resolution of better than 200 microm and high-sensitivity detection of 580-microm blood tubing to depths greater than 3 cm in a turbid medium with reduced scattering coefficient mu(s) (')=7.8 cm(-1). The system is used to systematically investigate the effects of target size, orientation, and geometry on tomographic imaging. As a demonstration of these effects and the system imaging capabilities, we present tomographic photoacoustic images of the brain vasculature of an ex vivo mouse with varying measurement aperture. For the first time, according to our knowledge, resolution of sub-200-microm vessels with an overlying turbid medium of greater than 2 cm depth is demonstrated using only intrinsic biological contrast.

Image Distortion in Thermoacoustic Tomography Caused by Microwave Diffraction

Physical Review. E, Statistical, Nonlinear, and Soft Matter Physics. Mar, 2008  |  Pubmed ID: 18517438

We report an intrinsic image distortion in microwave-induced thermoacoustic tomography. The distortion, due to microwave diffraction in the object to be imaged, leads to nonuniform excitation of acoustic pressure during microwave illumination. Both numerical simulations and phantom experiments demonstrate this phenomenon. A method of partial correction is also provided.

Realtime Photoacoustic Microscopy in Vivo with a 30-MHz Ultrasound Array Transducer

Optics Express. May, 2008  |  Pubmed ID: 18545502

We present a novel high-frequency photoacoustic microscopy system capable of imaging the microvasculature of living subjects in realtime to depths of a few mm. The system consists of a high-repetition-rate Q-switched pump laser, a tunable dye laser, a 30-MHz linear ultrasound array transducer, a multichannel high-frequency data acquisition system, and a shared-RAM multi-core-processor computer. Data acquisition, beamforming, scan conversion, and display are implemented in realtime at 50 frames per second. Clearly resolvable images of 6-microm-diameter carbon fibers are experimentally demonstrated at 80 microm separation distances. Realtime imaging performance is demonstrated on phantoms and in vivo with absorbing structures identified to depths of 2.5-3 mm. This work represents the first high-frequency realtime photoacoustic imaging system to our knowledge.

In Vivo Burn Imaging Using Mueller Optical Coherence Tomography

Optics Express. Jul, 2008  |  Pubmed ID: 18607436

We report on the use of a high-speed, fiber-based Mueller-matrix optical coherence tomography system with continuous source-polarization modulation for in vivo burn depth evaluation and healing monitoring. A homemade hand-held probe with integrated optical scanning and beam delivering optics was coupled in the sample arm. In vivo burn imaging was demonstrated on porcine skin at different stages of the wound healing process, where porcine skin was used because of its similarity to the human skin. Thermally damaged region was clearly localized in the depth-resolved phase retardation image extracted from the measured Jones matrix. The burn areas in the OCT images agreed well with the histology. By using a decomposition algorithm developed by our group, we also mapped the local birefringence of the skin. The experimental results demonstrate the system's potential for in vivo burn-depth determination.

Design and Evaluation of a Novel Breast Cancer Detection System Combining Both Thermoacoustic (TA) and Photoacoustic (PA) Tomography

Medical Physics. Jun, 2008  |  Pubmed ID: 18649451

We have developed a novel scanner for breast cancer detection, integrating both thermoacoustic and photoacoustic techniques to achieve dual contrast (microwave and light absorption) imaging. This scanner is nonionizing, low cost, and can potentially provide high-resolution, dual modality three-dimensional images of the breast. The scanner uses front instead of side breast compression and dry instead of gel ultrasonic coupling. Here we present the design of the breast scanner along with initial tissue phantom study results as a precursor to an actual patient study.

Effects of Acoustic Heterogeneities on Transcranial Brain Imaging with Microwave-induced Thermoacoustic Tomography

Medical Physics. Jul, 2008  |  Pubmed ID: 18697545

The effects of acoustic heterogeneities on transcranial brain imaging with microwave-induced thermoacoustic tomography were studied. A numerical model for calculating the propagation of thermoacoustic waves through the skull was developed and experimentally examined. The model takes into account wave reflection and refraction at the skull surfaces and therefore provides improved accuracy for the reconstruction. To evaluate when the skull-induced effects could be ignored in reconstruction, the reconstructed images obtained by the proposed method were further compared with those obtained with the method based on homogeneous acoustic properties. From simulation and experimental results, it was found that when the target region is close to the center of the brain, the effects caused by the skull layer are minimal and both reconstruction methods work well. As the target region becomes closer to the interface between the skull and brain tissue, however, the skull-induced distortion becomes increasingly severe, and the reconstructed image would be strongly distorted without correcting those effects. In this case, the proposed numerical method can improve image quality by taking into consideration the wave refraction and mode conversion at the skull surfaces. This work is important for obtaining good brain images when the thickness of the skull cannot be ignored.

Detection of Ultrasound-modulated Diffuse Photons Using Spectral-hole Burning

Optics Express. Sep, 2008  |  Pubmed ID: 18795023

The lack of efficient detection techniques has so far prevented ultrasound-modulated optical tomography from achieving maturity. By applying a quantum spectral filter based on spectral-hole burning, one modulation sideband of the ultrasound-modulated diffuse photons can be efficiently selected while the DC and the other sidebands are blocked. This technique features a large etendue as well as the capability of processing numerous speckles in parallel. It is also immune to speckle decorrelation, potentially allowing real-time in vivo imaging. Both theory and experiments are presented.

High-numerical-aperture-based Virtual Point Detectors for Photoacoustic Tomography

Applied Physics Letters. Jul, 2008  |  Pubmed ID: 18802493

The focal point of a high-numerical-aperture (NA) ultrasonic transducer can be used as a virtual point detector. This virtual point detector detects omnidirectionally over a wide acceptance angle. It also combines a large active transducer surface and a small effective virtual detector size. Thus the sensitivity is high compared with that of a real point detector, and the aperture effect is small compared with that of a finite size transducer. We present two kinds of high-NA-based virtual point detectors and their successful application in photoacoustic tomography. They can also be applied in other ultrasound-related fields.

Negative Lens Concept for Photoacoustic Tomography

Physical Review. E, Statistical, Nonlinear, and Soft Matter Physics. Aug, 2008  |  Pubmed ID: 18850859

Although a small point ultrasound transducer has a wide acceptance angle, its small active area leads to a high thermal-noise-induced electric voltage in the transducer, thus the sensitivity is low. By contrast, a finite-size flat transducer has high sensitivity, but the acceptance angle is small, which limits its application in reconstruction-based photoacoustic tomography (PAT). Here, we propose a negative lens concept to increase the acceptance angle of a flat transducer without losing sensitivity. Phantom experiments demonstrate that use of this concept greatly increases the detection region for PAT with high sensitivity.

Noninvasive Photoacoustic Imaging of the Thoracic Cavity and the Kidney in Small and Large Animals

Medical Physics. Oct, 2008  |  Pubmed ID: 18975699

The internal organs of rats and rabbits were clearly imaged noninvasively using a deeply penetrating reflection-mode photoacoustic imaging system. This imaging system had previously been found to provide an imaging depth limit of approximately 38 mm. In the thoracic cavity, major blood vessels connecting to the heart were imaged, and the right atrium was imaged as deeply as approximately 8 mm. In the abdominal cavities, the kidney and vena cava inferior were also imaged in situ. The vena cava inferior approximately 13.7 mm deep was imaged. The kidney of a New Zealand white rabbit was also imaged. This study shows the deep internal organ imaging capability of the system in animals. This technology can potentially be used to study tumors in internal organs, and be adapted to clinical diagnosis.

Monkey Brain Cortex Imaging by Photoacoustic Tomography

Journal of Biomedical Optics. Jul-Aug, 2008  |  Pubmed ID: 19021337

Photoacoustic tomography (PAT) is applied to image the brain cortex of a monkey through the intact scalp and skull ex vivo. The reconstructed PAT image shows the major blood vessels on the monkey brain cortex. For comparison, the brain cortex is imaged without the scalp, and then imaged again without the scalp and skull. Ultrasound attenuation through the skull is also measured at various incidence angles. This study demonstrates that PAT of the brain cortex is capable of surviving the ultrasound signal attenuation and distortion caused by a relatively thick skull.

Fast 3-D Dark-field Reflection-mode Photoacoustic Microscopy in Vivo with a 30-MHz Ultrasound Linear Array

Journal of Biomedical Optics. Sep-Oct, 2008  |  Pubmed ID: 19021408

We present an in vivo dark-field reflection-mode photoacoustic microscopy system that performs cross-sectional (B-scan) imaging at 50 Hz with real-time beamforming and 3-D imaging consisting of 166 B-scan frames at 1 Hz with postbeamforming. To our knowledge, this speed is currently the fastest in photoacoustic imaging. A custom-designed light delivery system is integrated with a 30-MHz ultrasound linear array to realize dark-field reflection-mode imaging. Linear mechanical scanning of the array produces 3-D images. The system has axial, lateral, and elevational resolutions of 25, 70, and 200 microm, respectively, and can image 3 mm deep in scattering biological tissues. Volumetric images of subcutaneous vasculature in rats are demonstrated in vivo. Fast 3-D photoacoustic microscopy is anticipated to facilitate applications of photoacoustic imaging in biomedical studies that involve dynamics and clinical procedures that demand immediate diagnosis.

Noninvasive Photoacoustic Identification of Sentinel Lymph Nodes Containing Methylene Blue in Vivo in a Rat Model

Journal of Biomedical Optics. Sep-Oct, 2008  |  Pubmed ID: 19021413

Sentinel lymph node biopsy (SLNB) has become the standard method of axillary staging for patients with breast cancer and clinically negative axillae. Even though SLNB using both methylene blue and radioactive tracers has a high identification rate, it still relies on an invasive surgical procedure with associated morbidity. Axillary ultrasound has emerged as a diagnostic tool to evaluate the axilla, but it can only assess morphology and cannot specifically identify sentinel lymph nodes (SLNs). In this pilot study, we propose a noninvasive photoacoustic SLN identification system using methylene blue injection in a rat model. We successfully image a SLN with high optical contrast (146+/-41, standard deviation) and good ultrasonic resolution (approximately 500 microm) in vivo. We also show potential feasibility for clinical applications by imaging 20- and 31-mm-deep SLNs in 3-D and 2-D, respectively. Our results suggest that this technology would be a useful clinical tool, allowing clinicians to identify SLNs noninvasively in vivo.

Ultrasound-modulated Optical Microscopy

Journal of Biomedical Optics. Sep-Oct, 2008  |  Pubmed ID: 19021426

We demonstrate that microscopic imaging is feasible in ultrasound-modulated optical tomography (UOT) of soft biological tissues, using a high-frequency focused ultrasound transducer with a 75-MHz central frequency. Our experiments in tissue mimicking phantoms show that at an imaging depth of about 2 mm, an axial resolution better than 30 microm can be achieved, whereas the lateral resolution is 38 microm. A long-cavity scanning confocal Fabry-Perot interferometer (CFPI) is used for real-time detection of multiply scattered light modulated by high-frequency ultrasound pulses propagating in an optically scattering medium. We also compare the performances of various high-frequency focused ultrasound transducers with central frequencies of 15 MHz, 30 MHz, 50 MHz, and 75 MHz. The comparison is based on two-dimensional (2-D) images of optically absorbing objects positioned at a few millimeters depth below the surface of both optically scattering phantoms and soft biological tissue samples. Our experimental results show that modulation depth and image contrast decrease with an increase in ultrasound frequency. In addition, we use analytical calculations to show that modulation depth decreases with increasing ultrasound frequency.

Pulsed Ultrasound-modulated Optical Tomography Using Spectral-hole Burning As a Narrowband Spectral Filter

Applied Physics Letters. Jul, 2008  |  Pubmed ID: 19079748

We applied a submegahertz nonlinear optical filter afforded by a cryogenically cooled spectral-hole burning crystal to ultrasound-modulated optical tomography. Our experimental results show that this technique, having the largest etendue among all available ultrasound-modulated optical tomography techniques and being immune to speckle decorrelation, offers potential for imaging in vivo and forming high resolution optical tomograms in real time. It opens an opportunity for the development of a clinically applicable high resolution optical imaging modality.

Adaptive and Robust Methods of Reconstruction (ARMOR) for Thermoacoustic Tomography

IEEE Transactions on Bio-medical Engineering. Dec, 2008  |  Pubmed ID: 19126453

In this paper, we present new adaptive and robust methods of reconstruction (ARMOR) for thermoacoustic tomography (TAT), and study their performances for breast cancer detection. TAT is an emerging medical imaging technique that combines the merits of high contrast due to electromagnetic or laser stimulation and high resolution offered by thermal acoustic imaging. The current image reconstruction methods used for TAT, such as the delay-and-sum (DAS) approach, are data-independent and suffer from low-resolution, high sidelobe levels, and poor interference rejection capabilities. The data-adaptive ARMOR can have much better resolution and much better interference rejection capabilities than their data-independent counterparts. By allowing certain uncertainties, ARMOR can be used to mitigate the amplitude and phase distortion problems encountered in TAT. The excellent performance of ARMOR is demonstrated using both simulated and experimentally measured data.

A 3-D High-frequency Array Based 16 Channel Photoacoustic Microscopy System for in Vivo Micro-vascular Imaging

IEEE Transactions on Medical Imaging. Aug, 2009  |  Pubmed ID: 19131292

This paper discusses the design of a novel photoacoustic microscopy imaging system with promise for studying the structure of tissue microvasculature for applications in visualizing angiogenesis. A new 16 channel analog and digital high-frequency array based photoacoustic microscopy system (PAM) was developed using an Nd:YLF pumped tunable dye laser, a 30 MHz piezo composite linear array transducer, and a custom multichannel receiver electronics system. Using offline delay and sum beamforming and beamsteering, phantom images were obtained from a 6 mum carbon fiber in water at a depth of 8 mm. The measured -6 dB lateral and axial spatial resolution of the system was 100+/-5 microm and 45+/-5 microm, respectively. The dynamic focusing capability of the system was demonstrated by imaging a composite carbon fiber matrix through a 12.5 mm imaging depth. Next, 2-D in vivo images were formed of vessels around 100 mum in diameter in the human hand. Three-dimensional in vivo images were also formed of micro-vessels 3 mm below the surface of the skin in two Sprague Dawley rats.

Near-infrared Gold Nanocages As a New Class of Tracers for Photoacoustic Sentinel Lymph Node Mapping on a Rat Model

Nano Letters. Jan, 2009  |  Pubmed ID: 19072058

This work demonstrated the use of Au nanocages as a new class of lymph node tracers for noninvasive photoacoustic (PA) imaging of a sentinel lymph node (SLN). Current SLN mapping methods based on blue dye and/or nanometer-sized radioactive colloid injection are intraoperative due to the need for visual detection of the blue dye and low spatial resolution of Geiger counters in detecting radioactive colloids. Compared to the current methods, PA mapping based on Au nanocages shows a number of attractive features: noninvasiveness, strong optical absorption in the near-infrared region (for deep penetration), and the accumulation of Au nanocages with a higher concentration than the initial solution for the injection. In an animal model, these features allowed us to identify SLNs containing Au nanocages as deep as 33 mm below the skin surface with good contrast. Most importantly, compared to methylene blue Au nanocages can be easily bioconjugated with antibodies for targeting specific receptors, potentially eliminating the need for invasive axillary staging procedures in addition to providing noninvasive SLN mapping.

M-mode Photoacoustic Particle Flow Imaging

Optics Letters. Mar, 2009  |  Pubmed ID: 19252588

Recently, there has been growing interest in the development of photoacoustic flow measuring methods aimed to study microvascular blood flow in biological tissue. Here, we describe the M-mode photoacoustic particle flow imaging, using an optical resolution photoacoustic microscope equipped with a high-repetition-rate pulsed dye laser. We studied the flow of a diluted dyed particle suspension in a small tube embedded in a nonscattering medium as well as in a scattering medium simulating biological tissue. Potentially, the method can be applied to detect the flow speed of single red blood cells in a capillary.

In-vivo Photoacoustic Microscopy of Nanoshell Extravasation from Solid Tumor Vasculature

Journal of Biomedical Optics. Jan-Feb, 2009  |  Pubmed ID: 19256687

In this study, high resolution backward-mode photoacoustic microscopy (PAM) is used to noninvasively image progressive extravasation and accumulation of nanoshells within a solid tumor in vivo. PAM takes advantage of the strong near-infrared absorption of nanoshells and their extravasation tendency from leaky tumor vasculatures for imaging. Subcutaneous tumors are grown on immunocompetent BALB/c mice. Polyethylene glycol (PEGylated) nanoshells with a peak optical absorption at approximately 800 nm are intravenously administered. With an 800-nm laser source, a prescan prior to nanoshell injection is taken to determine the background that is free of nanoshell accumulation. After injection, the 3-D nanoshell distribution at the tumor foci is monitored by PAM for 6 h. Experimental results show that accumulated nanoshells delineate the tumor position. Nanoshell accumulation is heterogeneous in tumors: more concentrated within the tumor cortex and largely absent from the tumor core. Because nanoshells have been recently demonstrated to enhance thermal therapy of subcutaneous tumors, we anticipate that PAM will be an important aid before, during, and after nanoshell thermal therapy.

Ex Vivo Blood Vessel Imaging Using Ultrasound-modulated Optical Microscopy

Journal of Biomedical Optics. Jan-Feb, 2009  |  Pubmed ID: 19256703

Recently we developed ultrasound-modulated optical microscopy (UOM) based on a long-cavity confocal Fabry-Perot interferometer (CFPI). This interferometer is used for real-time detection of multiply scattered light modulated by high frequency (30 to 75 MHz) ultrasound pulses propagating in an optically, strongly scattering medium. In this work, we use this microscope to study the dependence of ultrasound-modulated optical signals on the optical absorption and scattering properties of objects embedded about 3 mm deep in tissue mimicking phantoms. These results demonstrate that UOM has the potential to map both optical absorption and scattering contrast. Most importantly, for the first time in the field of ultrasound-modulated optical imaging, we image blood vasculature in highly scattering tissue samples from a mouse and a rat. Therefore, UOM could be a promising tool to study the morphology of blood vasculature and blood-associated functional parameters, such as oxygen saturation.

Noninvasive in Vivo Spectroscopic Nanorod-contrast Photoacoustic Mapping of Sentinel Lymph Nodes

European Journal of Radiology. May, 2009  |  Pubmed ID: 19269762

Sentinel lymph node (SLN) biopsy has increasingly become important in axillary staging of breast cancer patients since SLN biopsy alleviates the postoperative complications of previously practiced axillary lymph node dissections. Nevertheless, the procedures of SLN biopsy using blue dye and radioactive substance are still intraoperative, and the latter methods are also ionizing. In this pilot study, we have proposed noninvasive in vivo spectroscopic photoacoustic (PA) SLN mapping using gold nanorods as lymph node tracers in a rat model. Gold nanorods have biocompatibility, high optical absorption, and easily tuned surface plasmon resonance peak wavelength.

Noninvasive Label-free Imaging of Microhemodynamics by Optical-resolution Photoacoustic Microscopy

Optics Express. Apr, 2009  |  Pubmed ID: 19399148

In vivo microcirculatory imaging facilitates the fundamental understanding of many major diseases. However, existing techniques generally require invasive procedures or exogenous contrast agents, which perturb the intrinsic physiology of the microcirculation. Here, we report on optical-resolution photoacoustic microscopy (OR-PAM) for noninvasive label-free microcirculatory imaging at cellular levels. For the first time, OR-PAM demonstrates quantification of hemoglobin concentration and oxygenation in single microvessels down to capillaries. Using this technique, we imaged several important yet elusive microhemodynamic activities-including vasomotion and vasodilation-in small animals in vivo. OR-PAM enables functional volumetric imaging of the intact microcirculation, thereby providing greatly improved accuracy and versatility for broad biological and clinical applications.

Noninvasive, in Vivo Imaging of Blood-oxygenation Dynamics Within the Mouse Brain Using Photoacoustic Microscopy

Journal of Biomedical Optics. Mar-Apr, 2009  |  Pubmed ID: 19405708

Photoacoustic microscopy (PAM) has been used to obtain high-resolution, noninvasive images of the in vivo mouse brain. In this work, we exploit the high-depth and temporal resolutions of PAM to noninvasively image the blood-oxygenation dynamics of multiple cortex vessels in the mouse brain simultaneously in response to controlled hypoxic and hyperoxic challenges. These results confirm the ability of PAM to track blood oxygenation in the mouse brain, a critical aspect of imaging brain activity through the hemodynamic response.

Ultrasound-modulated Optical Tomography in Reflection Mode with Ring-shaped Light Illumination

Journal of Biomedical Optics. Mar-Apr, 2009  |  Pubmed ID: 19405745

We have succeeded in implementing ring-shaped light illumination ultrasound-modulated optical tomography (UOT) in reflection mode. The system used intense acoustic bursts and a charge-coupled device (CCD) camera-based speckle contrast detection method. In addition, the implementation allows placing the tissue sample below (not within) an acoustic coupling water tank and scanning the tissue without moving the sample. Thus, the UOT system is more clinically applicable than previous transmission-mode systems. Furthermore, we have successfully imaged an ex vivo methylene-blue-dyed sentinel lymph node (SLN) embedded at a depth of 13 mm in chicken breast tissue. This UOT system offers several advantages: noninvasiveness, nonionizing radiation, portability, cost effectiveness, and the possibility of combination with ultrasound pulse-echo imaging and photoacoustic imaging. One potential application of the UOT system is mapping SLNs in axillary staging for breast cancer patients.

Tangential Resolution Improvement in Thermoacoustic and Photoacoustic Tomography Using a Negative Acoustic Lens

Journal of Biomedical Optics. Mar-Apr, 2009  |  Pubmed ID: 19405757

We developed a novel concept of using a negative acoustic lens to increase the acceptance angle of an unfocused large-area ultrasonic transducer (detector), leading to more than twofold improvement of the tangential resolution in both thermoacoustic and photoacoustic tomography. In both thermoacoustic and photoacoustic tomography, for a given transducer bandwidth, the aperture size of the detector affects the tangential resolution greatly when the object of interest is near the detector surface. We were able to overcome such tangential resolution deterioration by attaching an acoustic concave lens, made of acrylic in front of the flat detector surface. We then quantified the tangential resolution improvement using phantom images. We also showed that the use of the negative lens preserves the shape of an object after the image is reconstructed.

Photoacoustic Tomography of the Mouse Cerebral Cortex with a High-numerical-aperture-based Virtual Point Detector

Journal of Biomedical Optics. Mar-Apr, 2009  |  Pubmed ID: 19405775

The mouse cerebral cortex was imaged in situ by photoacoustic tomography (PAT). Instead of a flat ultrasonic transducer, a virtual point detector based on a high numerical aperture (NA), positively focused transducer was used. This virtual point detector has a wide omnidirectional acceptance angle, a high sensitivity, and a negligible aperture effect. In addition, the virtual point detector can be located much more closely to the object during the detection. Compared with a finite-size flat transducer, images generated by using this virtual point detector have both uniform signal-to-noise ratio (SNR) and resolution.

Automatic Algorithm for Skin Profile Detection in Photoacoustic Microscopy

Journal of Biomedical Optics. Mar-Apr, 2009  |  Pubmed ID: 19405778

We have developed an automatic algorithm to detect the skin profiles in the volumetric data acquired by photoacoustic microscopy for subcutaneous vasculature imaging. This algorithm analyzes the relationship between amplitudes of photoacoustic signals generated from the skin surface and underlying blood vessels to achieve a rough estimation of the skin profile. A better approximation of the skin profile is then acquired after nonparametric smoothing and Gaussian low-pass spatial filtering. An auto-fit scan mechanism is further developed based on the detected skin profile to achieve good ultrasonic focusing on the subcutaneous vessel layer when the skin contour variation is much larger than the ultrasonic focal zone. The importance of skin profile detection in calculating the maximum-amplitude-projection images and significantly improving the image quality by employing the auto-fit scan are demonstrated by in vivo experimental results.

Molecular Photoacoustic Tomography with Colloidal Nanobeacons

Angewandte Chemie (International Ed. in English). 2009  |  Pubmed ID: 19418503

Spotting clots: Vascularly constrained colloidal gold nanobeacons (GNBs; see picture) can be used as exogenous photoacoustic contrast agents for the targeted detection of fibrin, a major biochemical feature of thrombus. Fibrin-targeted GNBs provide a more than tenfold signal enhancement in photoacoustic tomography in the near-IR wavelength window, indicating their potential for diagnostic imaging.

In Vivo Carbon Nanotube-enhanced Non-invasive Photoacoustic Mapping of the Sentinel Lymph Node

Physics in Medicine and Biology. Jun, 2009  |  Pubmed ID: 19430111

Sentinel lymph node biopsy (SLNB), a less invasive alternative to axillary lymph node dissection (ALND), has become the standard of care for patients with clinically node-negative breast cancer. In SLNB, lymphatic mapping with radio-labeled sulfur colloid and/or blue dye helps identify the sentinel lymph node (SLN), which is most likely to contain metastatic breast cancer. Even though SLNB, using both methylene blue and radioactive tracers, has a high identification rate, it still relies on an invasive surgical procedure, with associated morbidity. In this study, we have demonstrated a non-invasive single-walled carbon nanotube (SWNT)-enhanced photoacoustic (PA) identification of SLN in a rat model. We have successfully imaged the SLN in vivo by PA imaging (793 nm laser source, 5 MHz ultrasonic detector) with high contrast-to-noise ratio (=89) and good resolution ( approximately 500 microm). The SWNTs also show a wideband optical absorption, generating PA signals over an excitation wavelength range of 740-820 nm. Thus, by varying the incident light wavelength to the near infrared region, where biological tissues (hemoglobin, tissue pigments, lipids and water) show low light absorption, the imaging depth is maximized. In the future, functionalization of the SWNTs with targeting groups should allow the molecular imaging of breast cancer.

Photoacoustic Endoscopy

Optics Letters. May, 2009  |  Pubmed ID: 19448831

We have developed photoacoustic endoscopy with a miniaturized imaging probe. A light-guiding optical fiber, an ultrasonic sensor, and a mechanical scanning unit are integrated to enable circumferential sector scanning, which produces B-scan images. Biological tissues, including the gastrointestinal tract of a rat, have been imaged ex vivo or in situ.

Optical Fluence Distribution Study in Tissue in Dark-field Confocal Photoacoustic Microscopy Using a Modified Monte Carlo Convolution Method

Applied Optics. Jun, 2009  |  Pubmed ID: 19516366

We have modified the existing convolution method of the Monte Carlo simulation for finite photon beams with both translational and rotational invariance. The modified convolution method was applied to simulate the optical fluence distribution in tissue in dark-field confocal photoacoustic microscopy. We studied the influence of the size of the dark field and the illumination incident angle on the depth position of the effective optical focus (the region with the highest fluence) and the fluence ratio (the ratio of the optical fluence at the effective optical focus inside the tissue to the optical fluence on the tissue surface along the ultrasonic axis). Within the reach of diffuse photons, the depth position of the effective optical focus increases with the size of the dark field and is much less sensitive to the incident angle. The findings show that, while the fluence at the effective optical focus decreases, the fluence ratio increases with the size of the dark field. The incident angle has a weaker influence on the fluence ratio than the size of the dark field does. An incident angle between 30 and 50 degrees gives the highest fluence at the effective optical focus.

Nanoparticles for Photoacoustic Imaging

Wiley Interdisciplinary Reviews. Nanomedicine and Nanobiotechnology. Jul-Aug, 2009  |  Pubmed ID: 20049803

Nanoparticles have been designed and applied as contrast enhancers in various optical imaging techniques, such as optical coherence tomography, fluorescence imaging, and optical reflectance microscopy. As an emerging hybrid imaging modality, photoacoustic imaging (PAI) has also benefited from the application of these nanoparticle-based contrast agents. We review this rapidly growing field and describe the applications of nanoparticles in PAI. Particular focus is given to nanoparticles whose absorption mechanism is based on surface plasmon resonance, including gold nanoshells, nanorods, and nanocages. Dye-embedded nanoparticles are also reviewed. Specifically, the design and application of each nanoparticle-based contrast agent in relation to the field of PAI are detailed.

Multiscale Photoacoustic Microscopy and Computed Tomography

Nature Photonics. Aug, 2009  |  Pubmed ID: 20161535

Photoacoustic tomography (PAT) is probably the fastest growing biomedical imaging technology owing to its capability of high-resolution sensing of rich optical contrast in vivo at depths beyond the optical transport mean free path (~1 mm in the skin). Existing high-resolution optical imaging technologies, such as confocal microscopy and two-photon microscopy, have fundamentally impacted biomedicine but cannot reach such depths. Taking advantage of low ultrasonic scattering, PAT indirectly improves tissue transparency by 100 to 1000 fold and consequently enables deeply penetrating functional and molecular imaging at high spatial resolution. Further, PAT holds the promise of in vivo imaging at multiple length scales ranging from subcellular organelles to organs with the same contrast origin, an important application in multiscale systems biology research.

A Real-time Photoacoustic Tomography System for Small Animals

Optics Express. Jun, 2009  |  Pubmed ID: 19550444

A real-time 512-element photoacoustic tomography system for small animal imaging using a ring ultrasound array has been developed. The system, based upon a 5 MHz transducer array formed along a 50 mm circular aperture, achieves sub-200 micron lateral resolution over a 2 cm disk-shaped region. Corresponding elevation resolutions of 0.6 to 2.5 mm over the central volume enable depth-resolved 3D tomographic imaging with linear translation. Using 8:1 electronic multiplexing, imaging at up to 8 frame/sec is demonstrated for both dynamic phantoms and in vivo mouse and brain samples. The real-time, full 2D tomographic capability of the system paves the way for functional photoacoustic tomographic imaging studies in small animals with sub-second time frame.

Single-walled Carbon Nanotubes As a Multimodal-thermoacoustic and Photoacoustic-contrast Agent

Journal of Biomedical Optics. May-Jun, 2009  |  Pubmed ID: 19566311

We have developed a novel carbon nanotube-based contrast agent for both thermoacoustic and photoacoustic tomography. In comparison to deionized water, single-walled carbon nanotubes exhibited more than twofold signal enhancement for thermoacoustic tomography at 3 GHz. In comparison to blood, they exhibited more than sixfold signal enhancement for photoacoustic tomography at 1064 nm wavelength. The large contrast enhancement of single-walled carbon nanotubes was further corroborated by tissue phantom imaging studies.

In Vivo Functional Chronic Imaging of a Small Animal Model Using Optical-resolution Photoacoustic Microscopy

Medical Physics. Jun, 2009  |  Pubmed ID: 19610320

Optical-resolution photoacoustic microscopy (OR-PAM) has been validated as a valuable tool for label-free volumetric microvascular imaging. More importantly, the advantages of noninvasiveness and measurement consistency suggest the use of OR-PAM for chronic imaging of intact microcirculation. Here, such chronic imaging is demonstrated for the first time by monitoring the healing process of laser-induced microvascular lesions in a small animal model in vivo. The central part of a 1 mm by 1 mm region in a nude mouse ear was treated under a continuous-wave laser to create a microvascular lesion for chronic study. The region of interest was imaged before the laser treatment, immediately after the treatment, and throughout the healing process using both the authors' OR-PAM system and a commercial transmission-mode optical microscope. Three-dimensional microvascular morphology and blood oxygenation information were imaged simultaneously at capillary-level resolution. Transmission-mode optical microscopic images were acquired for comparison. OR-PAM has potential important applications in microcirculatory physiology or pathophysiology, tumor angiogenesis, laser microsurgery, and neuroscience.

Noninvasive, in Vivo Imaging of the Mouse Brain Using Photoacoustic Microscopy

Journal of Applied Physics. May, 2009  |  Pubmed ID: 19657402

Noninvasive, high resolution imaging of mouse brain activity is poised to provide clinically translatable insights into human neurological disease progression. Toward noninvasive imaging of brain activity through the hemodynamic response, the dark-field photoacoustic microscopy (PAM) technique was enhanced to image the cortex vasculature of the mouse brain in vivo using endogenous hemoglobin contrast. Specifically, the PAM system was redesigned to efficiently collect photoacoustic waves originating from cortical vessels, providing high (70 mum lateral and 54 mum axial) resolution images of the mouse brain vasculature with a contrast-to-noise ratio of 25 dB. These findings confirm the efficacy of PAM to noninvasively image vascular structures in the mouse brain and the potential to image mouse brain function by tracking the hemodynamic response.

Measuring the Optical Absorption Cross-sections of Au-Ag Nanocages and Au Nanorods by Photoacoustic Imaging

The Journal of Physical Chemistry. C, Nanomaterials and Interfaces. May, 2009  |  Pubmed ID: 19680423

This paper presents a method for measuring the optical absorption cross-sections (σ(a)) of Au-Ag nanocages and Au nanorods. The method is based on photoacoustic (PA) imaging, where the detected signal is directly proportional to the absorption coefficient (μ(a)) of the nanostructure. For each type of nanostructure, we firstly obtained μ(a) from the PA signal by benchmarking against a linear calibration curve (PA signal vs. μ(a)) derived from a set of methylene blue solutions with different concentrations. We then calculated σ(a) by dividing the μ(a) by the corresponding concentration of the Au nanostructure. Additonally, we obtained the extinction cross-section (σ(e), sum of absorption and scattering) from the extinction spectrum recorded using a conventional UV-vis-NIR spectrometer. From the measurements of σ(a) and σ(e), we were able to easily derive both the absorption and scattering cross-sections for each type of gold nanostructure. The ratios of absorption to extinction obtained from experimental and theoretical approaches agreed well, demonstrating the potential use of this method in determining the optical absorption and scattering properties of gold nanostructures and other types of nanomaterials.

Photoacoustic Tomography and Sensing in Biomedicine

Physics in Medicine and Biology. Oct, 2009  |  Pubmed ID: 19724102

Photoacoustics has been broadly studied in biomedicine, for both human and small animal tissues. Photoacoustics uniquely combines the absorption contrast of light or radio frequency waves with ultrasound resolution. Moreover, it is non-ionizing and non-invasive, and is the fastest growing new biomedical method, with clinical applications on the way. This review provides a brief recap of recent developments in photoacoustics in biomedicine, from basic principles to applications. The emphasized areas include the new imaging modalities, hybrid detection methods, photoacoustic contrast agents and the photoacoustic Doppler effect, as well as translational research topics.

Functional Transcranial Brain Imaging by Optical-resolution Photoacoustic Microscopy

Journal of Biomedical Optics. Jul-Aug, 2009  |  Pubmed ID: 19725708

Optical-resolution photoacoustic microscopy (OR-PAM) is applied to functional brain imaging in living mice. A near-diffraction-limited bright-field optical illumination is employed to achieve micrometer lateral resolution, and a dual-wavelength measurement is utilized to extract the blood oxygenation information. The variation in hemoglobin oxygen saturation (sO(2)) along vascular branching has been imaged in a precapillary arteriolar tree and a postcapillary venular tree, respectively. To the best of our knowledge, this is the first report on in vivo volumetric imaging of brain microvascular morphology and oxygenation down to single capillaries through intact mouse skulls. It is anticipated that: (i) chronic imaging enabled by this minimally invasive procedure will advance the study of cortical plasticity and neurological diseases; (ii) revealing the neuroactivity-dependent changes in hemoglobin concentration and oxygenation will facilitate the understanding of neurovascular coupling at the capillary level; and (iii) combining functional OR-PAM and high-resolution blood flowmetry will have the potential to explore cellular pathways of brain energy metabolism.

High-speed Dynamic 3D Photoacoustic Imaging of Sentinel Lymph Node in a Murine Model Using an Ultrasound Array

Medical Physics. Aug, 2009  |  Pubmed ID: 19746805

Noninvasive photoacoustic sentinel lymph node (SLN) mapping with high spatial resolution has the potential to improve the false negative rate and eliminate the use of radioactive tracers in SLN identification. In addition, the demonstrated high spatial resolution may enable physicians to replace SLN biopsy with fine needle aspiration biopsy, and thus reduce the risk of associated morbidity. The primary goal of this study is to demonstrate the feasibility of high-speed 3D photoacoustic imaging of the uptake and clearance dynamics of Evans blue dye in SLNs. The photoacoustic imaging system was developed with a 30 MHz ultrasound array and a kHz repetition rate laser system. It acquires one 3D photoacoustic image of 166 B-scan frames in 1 s, with axial, lateral, and elevational resolutions of 25, 70, and 200 microm, respectively. With optic-fiber based light delivery, the entire system is compact and is convenient to use. Upon injection of Evans blue, a blue dye currently used in clinical SLN biopsy, SLNs in mice and rats were accurately and noninvasively mapped in vivo using our imaging system. In our experiments, the SLNs were found to be located at approximately 0.65 mm below the skin surface in mice and approximately 1.2 mm in rats. In some cases, lymph vessels and lymphatic valves were also imaged. The dye dynamics--accumulation and clearance--in SLNs were quantitatively monitored by sequential 3D imaging with temporal resolution of as high as approximately 6 s. The demonstrated capability suggests that high-speed 3D photoacoustic imaging should facilitate the understanding of the dynamics of various dyes in SLNs and potentially help identify SLNs with high accuracy.

Three-dimensional Combined Photoacoustic and Optical Coherence Microscopy for in Vivo Microcirculation Studies

Optics Express. Sep, 2009  |  Pubmed ID: 19770860

Photoacoustic microscopy is predominantly sensitive to optical absorption, while optical coherence tomography relies on optical backscattering. Integrating their complementary contrasts can provide comprehensive information about biological tissue. We have developed a dual-modality microscope that combines the two for studying microcirculation. Three-dimensional imaging of microvasculature and its local environment has been demonstrated at micrometer-order resolution using endogenous contrast in vivo.

On the Speckle-free Nature of Photoacoustic Tomography

Medical Physics. Sep, 2009  |  Pubmed ID: 19810480

A long-standing conundrum is why photoacoustic tomography (PAT) possesses the unique ability to produce images devoid of speckle artifacts while all other coherent imaging technologies do not.

Gold Nanocages Covered by Smart Polymers for Controlled Release with Near-infrared Light

Nature Materials. Dec, 2009  |  Pubmed ID: 19881498

Photosensitive caged compounds have enhanced our ability to address the complexity of biological systems by generating effectors with remarkable spatial/temporal resolutions. The caging effect is typically removed by photolysis with ultraviolet light to liberate the bioactive species. Although this technique has been successfully applied to many biological problems, it suffers from a number of intrinsic drawbacks. For example, it requires dedicated efforts to design and synthesize a precursor compound for each effector. The ultraviolet light may cause damage to biological samples and is suitable only for in vitro studies because of its quick attenuation in tissue. Here we address these issues by developing a platform based on the photothermal effect of gold nanocages. Gold nanocages represent a class of nanostructures with hollow interiors and porous walls. They can have strong absorption (for the photothermal effect) in the near-infrared while maintaining a compact size. When the surface of a gold nanocage is covered with a smart polymer, the pre-loaded effector can be released in a controllable fashion using a near-infrared laser. This system works well with various effectors without involving sophisticated syntheses, and is well suited for in vivo studies owing to the high transparency of soft tissue in the near-infrared region.

Effects of Different Imaging Models on Least-squares Image Reconstruction Accuracy in Photoacoustic Tomography

IEEE Transactions on Medical Imaging. Nov, 2009  |  Pubmed ID: 19884066

In the classic formulation of photoacoustic tomography (PAT), two distinct descriptions of the imaging model have been employed for developing reconstruction algorithms. We demonstrate that the numerical and statistical properties of unweighted least-squares reconstruction algorithms associated with each imaging model are generally very different. Specifically, some PAT reconstruction algorithms, including many of the iterative algorithms previously explored, do not work directly with the raw measured pressure wavefields, but rather with an integrated data function that is obtained by temporally integrating the photoacoustic wavefield. The integration modifies the statistical distribution of the data, introducing statistical correlations among samples. This change is highly significant for iterative algorithms, many of which explicitly or implicitly seek to minimize a statistical cost function. In this work, we demonstrate that iterative reconstruction by least-squares minimization yields better resolution-noise tradeoffs when working with the raw pressure data than with the integrated data commonly employed. In addition, we demonstrate that the raw-data based approach is less sensitive to certain deterministic errors, such as dc offset errors.

Photoacoustic Tomography of Small Animal Brain with a Curved Array Transducer

Journal of Biomedical Optics. Sep-Oct, 2009  |  Pubmed ID: 19895109

We present the application of a curved array photoacoustic tomographic imaging system that can provide rapid, high-resolution photoacoustic imaging of small animal brains. The system is optimized to produce a B-mode, 90-deg field-of-view image at sub-200-microm resolution at a frame rate of approximately 1 frame/second when a 10-Hz pulse repetition rate laser is employed. By rotating samples, a complete 360-deg scan can be achieved within 15 s. In previous work, two-dimensional (2-D) ex vivo mouse brain cortex imaging has been reported. We report three-dimensional (3-D) small animal brain imaging obtained with the curved array system. The results are presented as a series of 2-D cross-sectional images. Besides structural imaging, the blood oxygen saturation of the animal brain cortex is also measured in vivo. In addition, the system can measure the time-resolved relative changes in blood oxygen saturation level in the small animal brain cortex. Last, ultrasonic gel coupling, instead of the previously adopted water coupling, is conveniently used in near-real-time 2-D imaging.

Thermoacoustic and Photoacoustic Sensing of Temperature

Journal of Biomedical Optics. Sep-Oct, 2009  |  Pubmed ID: 19895126

We present a novel temperature-sensing technique using thermoacoustic and photoacoustic measurements. This noninvasive method has been demonstrated using a tissue phantom to have high temporal resolution and temperature sensitivity. Because both photoacoustic and thermoacoustic signal amplitudes depend on the temperature of the source object, the signal amplitudes can be used to monitor the temperature. A temperature sensitivity of 0.15 degrees C was obtained at a temporal resolution as short as 2 s, taking the average of 20 signals. The deep-tissue imaging capability of this technique can potentially lead us to in vivo temperature monitoring in thermal or cryogenic applications.

Evans Blue Dye-enhanced Capillary-resolution Photoacoustic Microscopy in Vivo

Journal of Biomedical Optics. Sep-Oct, 2009  |  Pubmed ID: 19895150

Complete and continuous imaging of microvascular networks is crucial for a wide variety of biomedical applications. Photoacoustic tomography can provide high resolution microvascular imaging using hemoglobin within red blood cells (RBCs) as an endogenic contrast agent. However, intermittent RBC flow in capillaries results in discontinuous and fragmentary capillary images. To overcome this problem, we use Evans blue (EB) dye as a contrast agent for in vivo photoacoustic imaging. EB has strong optical absorption and distributes uniformly in the blood stream by chemically binding to albumin. With the help of EB, complete and continuous microvascular networks--especially capillaries--are imaged. The diffusion dynamics of EB leaving the blood stream and the clearance dynamics of the EB-albumin complex are also quantitatively investigated.

High-efficiency and Side-viewing Micro Fiber Optic Probe for In-vivo Diffuse Reflectance Measurements of Human Epithelial Tissues

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: 19964486

In this paper, we present a new micro fiber optic probe for in-vivo diffuse reflectance measurements of human epithelial tissues. The probe consists of seven, 120 x 120 microm collection fibers and one 200 microm incidence source fiber, arranged into a side-viewing configuration by using curved air-core waveguides with high optical transmittance. The outer dimension of the probe is 2 x 8mm(2). The extremely small size, high transmission efficiency and side-viewing capability of the new probe make it suitable for endoscopic spectroscopic applications inside the human body.

Intravital Imaging of Amyloid Plaques in a Transgenic Mouse Model Using Optical-resolution Photoacoustic Microscopy

Optics Letters. Dec, 2009  |  Pubmed ID: 20016651

We report optical-resolution photoacoustic microscopy (OR-PAM) for in vivo imaging of amyloid plaques in an Alzheimer's disease mouse model. Validation using conventional fluorescence microscopy and multiphoton microscopy shows that OR-PAM has sufficient sensitivity and spatial resolution to identify amyloid plaques in living brains. In addition, with dual-wavelength OR-PAM, the three-dimensional morphology of amyloid plaques and the surrounding microvasculature are imaged simultaneously through a cranial window without angiographic contrast agents. OR-PAM, capable of providing both exogenous molecular contrast and endogenous hemoglobin contrast, has the potential to serve as a new technology for in vivo microscopic observations of cerebral plaque deposits.

Photoacoustic Imaging of Living Mouse Brain Vasculature Using Hollow Gold Nanospheres

Biomaterials. Mar, 2010  |  Pubmed ID: 20036000

Photoacoustic tomography (PAT) also referred to as optoacoustic tomography (OAT) is a hybrid imaging modality that employs nonionizing optical radiation and ultrasonic detection. Here, we describe the application of a new class of optical contrast agents based on mesoscopic hollow gold nanospheres (HAuNS) to PAT. HAuNS are approximately 40 nm in diameter with a hollow interior and consist of a thin gold wall. They display strong resonance absorption tuned to the near-infrared (NIR) range, with an absorption peak at 800 nm, whose photoacoustic efficiency is significantly greater than that of blood. Following surface conjugation with thiolated poly(ethylene glycol), the pegylated HAuNS (PEG-HAuNS) had distribution and elimination half-lives of 1.38 +/- 0.38 and 71.82 +/- 30.46 h, respectively. Compared with PAT images based on the intrinsic optical contrast in nude mice, the PAT images acquired within 2 h after intravenous administration of PEG-HAuNS showed the brain vasculature with greater clarity and detail. The image depicted brain blood vessels as small as approximately 100 mum in diameter using PEG-HAuNS as contrast agents. Preliminary results showed no acute toxicity to the liver, spleen, or kidneys in mice following a single imaging dose of PEG-HAuNS. Our results indicate that PEG-HAuNS are promising contrast agents for PAT, with high spatial resolution and enhanced sensitivity.

Real-time Photoacoustic Tomography of Cortical Hemodynamics in Small Animals

Journal of Biomedical Optics. Jan-Feb, 2010  |  Pubmed ID: 20210422

For the first time, the hemodynamics within the entire cerebral cortex of a mouse were studied by using photoacoustic tomography (PAT) in real time. The PAT system, based on a 512-element full-ring ultrasound array, received photoacoustic signals primarily from a slice of 2-mm thickness. This system can provide high-resolution brain vasculature images. We also monitored the fast wash-in process of a photoacoustic contrast agent in the mouse brain. Our results demonstrated that PAT is a powerful imaging modality that can be potentially used to study small animal neurofunctional activities.

Multifunctional Microbubbles and Nanobubbles for Photoacoustic and Ultrasound Imaging

Journal of Biomedical Optics. Jan-Feb, 2010  |  Pubmed ID: 20210423

We develop a novel dual-modal contrast agent-encapsulated-ink poly(lactic-co-glycolic acid) (PLGA) microbubbles and nanobubbles-for photoacoustic and ultrasound imaging. Soft gelatin phantoms with embedded tumor simulators of encapsulated-ink PLGA microbubbles and nanobubbles in various concentrations are clearly shown in both photoacoustic and ultrasound images. In addition, using photoacoustic imaging, we successfully image the samples positioned below 1.8-cm-thick chicken breast tissues. Potentially, simultaneous photoacoustic and ultrasound imaging enhanced by encapsulated-dye PLGA microbubbles or nanobubbles can be a valuable tool for intraoperative assessment of tumor boundaries and therapeutic margins.

Photoacoustic Imaging and Characterization of the Microvasculature

Journal of Biomedical Optics. Jan-Feb, 2010  |  Pubmed ID: 20210427

Photoacoustic (optoacoustic) tomography, combining optical absorption contrast and highly scalable spatial resolution (from micrometer optical resolution to millimeter acoustic resolution), has broken through the fundamental penetration limit of optical ballistic imaging modalities-including confocal microscopy, two-photon microscopy, and optical coherence tomography-and has achieved high spatial resolution at depths down to the diffusive regime. Optical absorption contrast is highly desirable for microvascular imaging and characterization because of the presence of endogenous strongly light-absorbing hemoglobin. We focus on the current state of microvascular imaging and characterization based on photoacoustics. We first review the three major embodiments of photoacoustic tomography: microscopy, computed tomography, and endoscopy. We then discuss the methods used to characterize important functional parameters, such as total hemoglobin concentration, hemoglobin oxygen saturation, and blood flow. Next, we highlight a few representative applications in microvascular-related physiological and pathophysiological research, including hemodynamic monitoring, chronic imaging, tumor-vascular interaction, and neurovascular coupling. Finally, several potential technical advances toward clinical applications are suggested, and a few technical challenges in contrast enhancement and fluence compensation are summarized.

Gold Nanocages for Cancer Imaging and Therapy

Methods in Molecular Biology (Clifton, N.J.). 2010  |  Pubmed ID: 20217590

Gold nanocages are hollow nanostructures with porous walls that can be simply prepared via the galvanic replacement reaction between silver nanocubes and chloroauric acid. Their optical resonance peaks can be precisely tuned into the near-infrared region, in which the adsorption caused by blood or soft tissue is essentially negligible. Significantly, the strong absorption of gold nanocages makes them attractive as a novel class of contrast enhancement and photothermal agents for cancer detection and treatment. The well-established chemistry for gold also allows them to target specific cells by functionalizing their surface with various moieties such as antibodies, peptides, and DNAs. In this chapter, we focus on their use as a photothermal agent for the ablation of cancer cells and as a contrast agent for the in vivo noninvasive photoacoustic imaging of blood vessels and the sentinel lymph nodes in rats.

Sentinel Lymph Nodes and Lymphatic Vessels: Noninvasive Dual-modality in Vivo Mapping by Using Indocyanine Green in Rats--volumetric Spectroscopic Photoacoustic Imaging and Planar Fluorescence Imaging

Radiology. May, 2010  |  Pubmed ID: 20413757

To noninvasively map sentinel lymph nodes (SLNs) and lymphatic vessels in rats in vivo by using dual-modality nonionizing imaging-volumetric spectroscopic photoacoustic imaging, which measures optical absorption, and planar fluorescence imaging, which measures fluorescent emission-of indocyanine green (ICG).

In Vivo Photoacoustic Imaging of Transverse Blood Flow by Using Doppler Broadening of Bandwidth

Optics Letters. May, 2010  |  Pubmed ID: 20436589

A method is proposed to measure transverse blood flow by using photoacoustic Doppler broadening of bandwidth. By measuring bovine blood flowing through a plastic tube, the linear dependence of the broadening on the flow speed was validated. The blood flow of the microvasculature in a mouse ear and a chicken embryo (stage 16) was also studied.

Section-illumination Photoacoustic Microscopy for Dynamic 3D Imaging of Microcirculation in Vivo

Optics Letters. May, 2010  |  Pubmed ID: 20436610

We developed section-illumination photoacoustic microscopy capable of dynamic in vivo imaging of microvessels as small as 30 microm in diameter. The section illumination improved the elevational resolution while an ultrasound array provided the in-plane axial and lateral resolutions. Using the imaging system, we monitored the wash-in dynamics of Evans Blue in the microcirculation of mouse ears at 249 Hz 2D and 0.5 Hz 3D image acquisition rates. Such observation allowed us to differentiate the arterioles from the venules. In the future, the technology may be used to study angiogenesis, diabetes-induced vascular complications, and pharmacokinetics.

Photoacoustic Microscopy with 2-microm Transverse Resolution

Journal of Biomedical Optics. Mar-Apr, 2010  |  Pubmed ID: 20459224

We present a new-generation optical-resolution confocal photoacoustic microscope, consisting of a 0.25-numerical aperture optical microscope objective and a 75-MHz center-frequency spherically focused ultrasonic transducer. Experiments verified that this microscope has a transverse resolution of 2 microm, which is the highest to our knowledge among all photoacoustic imaging systems. In situ imaging of mouse ears shows the feasibility of resolving individual red blood cells in microvessels using the current system.

Ultrasound-array-based Real-time Photoacoustic Microscopy of Human Pulsatile Dynamics in Vivo

Journal of Biomedical Optics. Mar-Apr, 2010  |  Pubmed ID: 20459225

With a refined ultrasound-array-based real-time photoacoustic microscopy (UA-PAM) system, we demonstrate the feasibility of noninvasive in vivo imaging of human pulsatile dynamics. The system, capable of real-time B-scan imaging at 50 Hz and high-speed 3-D imaging, is validated by imaging the subcutaneous microvasculature in rats and humans. After the validation, a human artery around the palm-wrist area is imaged, and its pulsatile dynamics, including the arterial pulsatile motion and changes in hemoglobin concentration, is monitored with 20-ms B-scan imaging temporal resolution. To our knowledge, this is the first demonstration of real-time photoacoustic imaging of human physiological dynamics. Our results show that UA-PAM can potentially enable many new possibilities for studying functional and physiological dynamics in both preclinical and clinical imaging settings.

Transverse Flow Imaging Based on Photoacoustic Doppler Bandwidth Broadening

Journal of Biomedical Optics. Mar-Apr, 2010  |  Pubmed ID: 20459226

We propose a new method to measure transverse flow velocity based on photoacoustic Doppler bandwidth broadening, which is determined by the geometry of the probe-beam and the velocity of the transverse flow. By exploiting pulsed laser excitation and raster motor scanning, three-dimensional structure and flow velocity can be imaged simultaneously. In addition, the flow direction can be determined with bidirectional scanning. In a flowing suspension of red-dyed microspheres (diameter: 6 microm), transverse flow speeds ranging from 0 to 2.5 mms as well as flow direction were measured. A cross-sectional flow image was also obtained with the tube laid in a zigzag pattern.

Compressed Sensing in Photoacoustic Tomography in Vivo

Journal of Biomedical Optics. Mar-Apr, 2010  |  Pubmed ID: 20459233

The data acquisition speed in photoacoustic computed tomography (PACT) is limited by the laser repetition rate and the number of parallel ultrasound detecting channels. Reconstructing an image with fewer measurements can effectively accelerate the data acquisition and reduce the system cost. We adapt compressed sensing (CS) for the reconstruction in PACT. CS-based PACT is implemented as a nonlinear conjugate gradient descent algorithm and tested with both phantom and in vivo experiments.

Saturation Effect in Functional Photoacoustic Imaging

Journal of Biomedical Optics. Mar-Apr, 2010  |  Pubmed ID: 20459239

We investigate the saturation effect, which describes the violation of the linearity between the measured photoacoustic amplitude and the object's optical absorption coefficient in functional photoacoustic imaging when the optical absorption in the object increases. We model the optical energy deposition and photoacoustic signal generation and detection in a semi-infinite optical absorbing object. Experiments are carried out by measuring photoacoustic signals generated from an ink-filled plastic tube. The saturation effect is studied by varying the optical absorption coefficient in the model and the ink concentration in the photoacoustic experiments. By changing the center frequency of the ultrasonic detector, the requirement to minimize the saturation effect in functional photoacoustic imaging is established.

Calibration-free Absolute Quantification of Optical Absorption Coefficients Using Acoustic Spectra in 3D Photoacoustic Microscopy of Biological Tissue

Optics Letters. Jun, 2010  |  Pubmed ID: 20548388

Optical absorption is closely associated with many physiological important parameters, such as the concentration and oxygen saturation of hemoglobin, and it can be used to quantify the concentrations of nonfluorescent molecules. We propose a method to use acoustic spectra of photoacoustic signals to quantify the absolute optical absorption. This method is self-calibrating and thus insensitive to variations in the optical fluence. Factors such as system bandwidth and acoustic attenuation can affect the quantification but can be canceled by dividing the acoustic spectra measured at two optical wavelengths. Using optical-resolution photoacoustic microscopy, we quantified the absolute optical absorption of black ink samples with various concentrations. We also quantified both the concentration and oxygen saturation of hemoglobin in a live mouse in absolute units.

Photoacoustic Tomography of Water in Phantoms and Tissue

Journal of Biomedical Optics. May-Jun, 2010  |  Pubmed ID: 20615021

Photoacoustic tomography (PAT) has been widely used to image optically absorptive objects in both human and animal tissues. For the first time, we present imaging of water with laser-based PAT. We photoacoustically measure the absorption spectra of water-ethanol mixtures at various water concentrations, and then image water-ethanol and pure-water inclusions in gel and a water inclusion in fat tissue. The significant difference in photoacoustic signals between water and fat tissue indicates that the laser-based PAT has the potential to detect water content in tissue.

Neurovascular Photoacoustic Tomography

Frontiers in Neuroenergetics. 2010  |  Pubmed ID: 20616885

Neurovascular coupling refers to the relationship between neuronal activities and downstream hemodynamic responses. Photoacoustic tomography (PAT), enabling comprehensive label-free imaging of hemodynamic activities with highly scalable penetration and spatial resolution, has great potential in the study of neurovascular coupling. In this review, we first introduce the technical basis of hemodynamic PAT - including label-free quantification of total hemoglobin concentration, blood oxygenation, and blood flow - as well as its applications in hemodynamic monitoring. Then, we demonstrate the potential application of PAT in neurovascular imaging by highlighting representative studies on cerebral vascular responses to whisker stimulation and Alzheimer's disease. Finally, potential research directions and associated technical challenges are discussed.

Integrated Photoacoustic and Fluorescence Confocal Microscopy

IEEE Transactions on Bio-medical Engineering. Oct, 2010  |  Pubmed ID: 20639165

We have developed a dual-modality imaging system by integrating optical-resolution photoacoustic microscopy and fluorescence confocal microscopy to provide optical absorption and fluorescence contrasts simultaneously. By sharing the same laser source and objective lens, intrinsically registered photoacoustic and fluorescence images are acquired in a single scan. The micrometer resolution allows imaging of both blood and lymphatic vessels down to the capillary level. Simultaneous photoacoustic angiography and fluorescence lymphangiography were demonstrated, presenting more information to study tumor angiogenesis, vasculature, and microenvironments in vivo.

Label-free Photoacoustic Ophthalmic Angiography

Optics Letters. Jan, 2010  |  Pubmed ID: 20664653

We present label-free functional photoacoustic imaging of the ocular microvasculature in living animals. The anterior segment of an adult mouse was imaged with a laser exposure level well within the American National Standards Institute safety standards. Individual red blood cells traveling along the iris capillaries were clearly resolved, and the hemoglobin oxygen saturation in the iris microvasculature was imaged spectrally. We believe that photoacoustic imaging has the potential to advance the diagnosis and treatment of ocular diseases in humans.

Chronic Label-free Volumetric Photoacoustic Microscopy of Melanoma Cells in Three-dimensional Porous Scaffolds

Biomaterials. Nov, 2010  |  Pubmed ID: 20727581

Visualizing cells in three-dimensional (3D) scaffolds has been one of the major challenges in tissue engineering. Most current imaging modalities either suffer from poor penetration depth or require exogenous contrast agents. Here, we demonstrate photoacoustic microscopy (PAM) of the spatial distribution and temporal proliferation of cells inside three-dimensional porous scaffolds with thicknesses over 1 mm. Specifically, we evaluated the effects of seeding and culture methods on the spatial distribution of melanoma cells. Spatial distribution of the cells in the scaffold was well-resolved in PAM images. Moreover, the number of cells in the scaffold was quantitatively measured from the as-obtained volumetric information. The cell proliferation profile obtained from PAM correlated well with what was obtained using the traditional 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.

In Vivo Molecular Photoacoustic Tomography of Melanomas Targeted by Bioconjugated Gold Nanocages

ACS Nano. Aug, 2010  |  Pubmed ID: 20731439

Early diagnosis, accurate staging, and image-guided resection of melanomas remain crucial clinical objectives for improving patient survival and treatment outcomes. Conventional techniques cannot meet this demand because of the low sensitivity, low specificity, poor spatial resolution, shallow penetration, and/or ionizing radiation. Here we overcome such limitations by combining high-resolution photoacoustic tomography (PAT) with extraordinarily optical absorbing gold nanocages (AuNCs). When bioconjugated with [Nle(4),D-Phe(7)]-alpha-melanocyte-stimulating hormone, the AuNCs can serve as a novel contrast agent for in vivo molecular PAT of melanomas with both exquisite sensitivity and high specificity. The bioconjugated AuNCs enhanced contrast approximately 300% more than the control, PEGylated AuNCs. The in vivo PAT quantification of the amount of AuNCs accumulated in melanomas was further validated with inductively coupled plasma mass spectrometry (ICP-MS).

Handheld Array-based Photoacoustic Probe for Guiding Needle Biopsy of Sentinel Lymph Nodes

Journal of Biomedical Optics. Jul-Aug, 2010  |  Pubmed ID: 20799812

By modifying a clinical ultrasound array system, we develop a novel handheld photoacoustic probe for image-guided needle biopsy. The integration of optical fiber bundles for pulsed laser light delivery enables photoacoustic image-guided insertion of a needle into rat axillary lymph nodes with accumulated indocyanine green (ICG). Strong photoacoustic contrast of the needle is achieved. After subcutaneous injection of the dye in the left forepaw, sentinel lymph nodes are easily detected, in vivo and in real time, beneath 2-cm-thick chicken breast overlaying the axillary region. ICG uptake in axillary lymph nodes is confirmed with fluorescence imaging both in vivo and ex vivo. These results demonstrate the clinical potential of this handheld photoacoustic system for facile identification and needle biopsy of sentinel lymph nodes for cancer staging and metastasis detection in humans.

Subwavelength-resolution Label-free Photoacoustic Microscopy of Optical Absorption in Vivo

Optics Letters. Oct, 2010  |  Pubmed ID: 20890331

Optical absorption provides essential biological functional information but cannot be sensed by mainstream optical microscopy technologies directly, which detect fluorescence or scattering and may require undesirable labeling. Here we developed in vivo subwavelength-resolution photoacoustic microscopy (SW-PAM) that provides exquisitely high optical-absorption contrast due to nonfluorescent, or fluorescent, endogenous pigments. Having approached the ultimate diffraction-limited optical resolution, SW-PAM can resolve subcellular organelles. Vasculature and early-stage melanoma were imaged with 12:1 and 17:1 contrasts, respectively, without labeling. SW-PAM along with the scaled-up macroscopy, as the only technology that measures the same contrast origin over such a wide length scale, can potentially accelerate translation from microscopic research to clinical practice.

Fast and Robust Deconvolution-based Image Reconstruction for Photoacoustic Tomography in Circular Geometry: Experimental Validation

IEEE Photonics Journal. Feb, 2010  |  Pubmed ID: 20953277

Photoacoustic tomography (PAT) is a fast-developing biomedical imaging technology suitable for in vivo imaging. PAT in spherical or circular geometry gives good image resolution yet is slow or expensive in signal acquisition and image formation. Reducing the number of detection angles can ameliorate such issues usually at the expense of image quality. This paper introduces a deconvolution-based algorithm that models the imaging process as a linear and shift-invariant system. As demonstrated by the in vivo experiment, this algorithm not only runs much faster than the back-projection algorithm, but also shows stronger robustness in that it provides better image quality when detection angles are sparse. Therefore, this algorithm promises to enable real-time PAT in circular geometry.

Picosecond Absorption Relaxation Measured with Nanosecond Laser Photoacoustics

Applied Physics Letters. Oct, 2010  |  Pubmed ID: 21079726

Picosecond absorption relaxation-central to many disciplines-is typically measured by ultrafast (femtosecond or picosecond) pump-probe techniques, which however are restricted to optically thin and weakly scattering materials or require artificial sample preparation. Here, we developed a reflection-mode relaxation photoacoustic microscope based on a nanosecond laser and measured picosecond absorption relaxation times. The relaxation times of oxygenated and deoxygenated hemoglobin molecules, both possessing extremely low fluorescence quantum yields, were measured at 576 nm. The added advantages in dispersion susceptibility, laser-wavelength availability, reflection sensing, and expense foster the study of natural-including strongly scattering and nonfluorescent-materials.

Near Infrared Photoacoustic Detection of Sentinel Lymph Nodes with Gold Nanobeacons

Biomaterials. May, 2010  |  Pubmed ID: 20172607

Detection of sentinel lymph node (SLN) using photoacoustic imaging is an emerging technique for noninvasive axillary staging of breast cancer. Due to the absence of intrinsic contrast inside the lymph nodes, exogenous contrast agents are used for photoacoustic detection. In this work, we have demonstrated near infrared detection of SLN with gold nanobeacons (GNBs) providing the photoacoustic contrast in a rodent model. We found that size dictates the in vivo characteristics of these nanoparticles in SLN imaging. Larger nanobeacons with high payloads of gold were not as efficient as smaller size nanobeacons with lower payloads for this purpose. Colloidal GNBs were designed as a nanomedicine platform with "soft" nature that is amenable to bio-elimination, an essential feature for in vivo efficacy and safety. The GNBs were synthesized as lipid- or polymer-encapsulated colloidal particles incorporating tiny gold nanoparticles (2-4 nm) in three tunable sizes (90 nm, 150 nm and 290 nm). Smaller GNBs were noted trafficking through the lymphatic system and accumulating more efficiently in the lymph nodes in comparison to the bigger nanoagents. At 20 min, the GNBs reached the SLN and were no longer observed within the draining lymphatic vessel. Within 1 h post-injection, the contrast ratio of the lymph nodes with the surrounding blood vessels was 9:1. These findings were also supported by analytical measurements of the ex vivo tissue samples. Results indicate that cumulative nanoparticle deposition in lymph nodes is size dependent and that high payloads of gold, although offering greater contrast in vitro, may yield nanoagents with poor intradermal migration and lymphatic transport characteristics.

In Vivo Photoacoustic Tomography of Chemicals: High-resolution Functional and Molecular Optical Imaging at New Depths

Chemical Reviews. May, 2010  |  Pubmed ID: 20210338

Special Section Guest Editorial: Photons Plus Ultrasound: Imaging and Sensing

Journal of Biomedical Optics. Mar-Apr, 2010  |  Pubmed ID: 20459223

Sentinel Lymph Nodes in the Rat: Noninvasive Photoacoustic and US Imaging with a Clinical US System

Radiology. Jul, 2010  |  Pubmed ID: 20574088

To evaluate in vivo sentinel lymph node (SLN) mapping by using photoacoustic and ultrasonographic (US) imaging with a modified clinical US imaging system.

A Facile Synthesis of Novel Self-assembled Gold Nanorods Designed for Near-infrared Imaging

Journal of Nanoscience and Nanotechnology. Dec, 2010  |  Pubmed ID: 21121304

Molecular imaging techniques now allow recognition of early biochemical, physiological, and anatomical changes before manifestation of gross pathological changes. Photoacoustic imaging represents a novel non-ionizing detection technique that combines the advantages of optical and ultrasound imaging. Noninvasive photoacoustic tomography (PAT) imaging in combination with nanoparticle-based contrast agents show promise in improved detection and diagnosis of cardiovascular and cancer related diseases. In this report, a novel strategy is introduced to achieve self-assembled colloidal gold nanorods, which are constrained to the vasculature. Gold nanorods (2-4 nm) were incorporated into the core of self-assembled lipid-encapsulated nanoparticles (sGNR) (approximately 130 nm), providing more than hundreds of gold atoms per nanoparticle of 20% colloid suspension. The physico-chemical characterization in solution and anhydrous state with analytical techniques demonstrated that the particles were spherical and highly mono dispersed. In addition to the synthesis and characterization, sensitive near-infrared photoacoustic detection was impressively demonstrated in vitro.

Photoacoustic Generation of Focused Quasi-unipolar Pressure Pulses

Journal of Innovative Optical Health Sciences. Jan, 2010  |  Pubmed ID: 21152369

The photoacoustic effect was employed to generate short-duration quasi-unipolar acoustic pressure pulses in both planar and spherically focused geometries. In the focal region, the temporal profile of a pressure pulse can be approximated by the first derivative of the temporal profile near the front transducer surface, with a time averaged value equal to zero. This approximation agreed with experimental results acquired from photoacoustic transducers with both rigid and free boundaries. For a free boundary, the acoustic pressure in the focal region is equal to the sum of a positive pressure that follows the spatial profile of the optical energy deposition in the medium and a negative pressure that follows the temporal profile of the laser pulse.

In Vivo Label-free Photoacoustic Microscopy of Cell Nuclei by Excitation of DNA and RNA

Optics Letters. Dec, 2010  |  Pubmed ID: 21165116

Imaging of cell nuclei plays a critical role in cancer diagnosis and prognosis. To image noninvasively cell nuclei in vivo without staining, we developed UV photoacoustic microscopy (UV-PAM), in which 266 nm wavelength UV light excites unlabeled DNA and RNA in cell nuclei to produce photoacoustic waves. We applied UV-PAM to ex vivo imaging of cell nuclei in a mouse lip and a mouse small intestine and to in vivo imaging of the cell nuclei in the mouse skin. The UV-PAM images of unstained cell nuclei match the optical micrographs of the histologically stained cell nuclei. Given intrinsic optical contrast and high spatial resolution, in vivo label-free UV-PAM has potential for unique biological and clinical application.

Spectral Hole Burning for Ultrasound-modulated Optical Tomography of Thick Tissue

Journal of Biomedical Optics. Nov-Dec, 2010  |  Pubmed ID: 21198192

We apply spectral hole burning (SHB)-aided detection in ultrasound-modulated optical tomography (UOT) to image optical heterogeneities in thick tissue-mimicking phantom samples and chicken breast tissue. The efficiency of SHB is improved by using a Tm(3+):YAG crystal of higher doping concentration (2.0-atomic%) and a double-pass pumping configuration, in which the pump beam is transmitted through the crystal twice to burn a deeper spectral hole with the available optical intensity. With the improved SHB-UOT system, we image absorbing, scattering, and phase objects that are embedded in the middle plane of a 30-mm-thick phantom sample. The imaging resolution was 0.5 mm in the lateral direction, as defined by the focal width of the ultrasonic transducer, and 1.5 mm in the axial direction, as determined by the ultrasonic burst length. We also image two absorbing objects embedded in a 32-mm-thick chicken breast sample. The results suggest that the improved SHB-UOT system is one step closer to the practical optical imaging application in biological and clinical studies.

Deeply Penetrating in Vivo Photoacoustic Imaging Using a Clinical Ultrasound Array System

Biomedical Optics Express. 2010  |  Pubmed ID: 21258465

Using a hand-held photoacoustic probe integrated with a clinical ultrasound array system, we successfully imaged objects deeply positioned in biological tissues. The optical contrasts were enhanced by methylene blue with a concentration of ~30 mM. The penetration depth reached ~5.2 cm in chicken breast tissue by using 650-nm wavelength, which is ~4.7 times the 1/e optical penetration depth. This imaging depth was achieved using a laser fluence on the tissue surface of only 3 mJ/cm(2), which is 1/7 of the American National Standards Institute (ANSI) safety limit (20 mJ/cm(2)). The noise equivalent sensitivity at this depth was ~11 mM. Further, after intradermal injection of methylene blue in a rat, a sentinel lymph node was easily detected in vivo, beneath a 2-cm thick layer of chicken breast. Also, blood located 3.5 cm deep in the rat was clearly imaged with intrinsic contrast. We have photoacoustically guided insertion of a needle into a rat sentinel lymph node with accumulated methylene blue. These results highlight the clinical potential of photoacoustic image-guided identification and needle biopsy of sentinel lymph nodes for axillary staging in breast cancer patients.

Fast Voice-coil Scanning Optical-resolution Photoacoustic Microscopy

Optics Letters. Jan, 2011  |  Pubmed ID: 21263479

We developed a photoacoustic imaging system that has real-time imaging capability with optical resolution. The imaging system is capable of scanning at 20 Hz over a 9 mm range and up to 40 Hz over a 1 mm scanning range. A focused laser beam provides a lateral resolution of 3.4 μm as measured in an optically nonscattering medium. Flows of micrometer-sized carbon particles or whole blood in a silicone tube and individual red blood cells (RBCs) in mouse ear capillaries were also imaged in real time, demonstrating the capability to image highly dynamic processes in vivo at a micrometer-scale resolution.

Fiber-laser-based Photoacoustic Microscopy and Melanoma Cell Detection

Journal of Biomedical Optics. Jan-Feb, 2011  |  Pubmed ID: 21280901

For broad applications in biomedical research involving functional dynamics and clinical studies, a photoacoustic microscopy system should be compact, stable, and fast. In this work, we use a fiber laser as the photoacoustic irradiation source to meet these goals. The laser system measures 45×56×13 cm3. The stability of the laser is attributed to the intrinsic optical fiber-based light amplification and output coupling. Its 50-kHz pulse repetition rate enables fast scanning or extensive signal averaging. At the laser wavelength of 1064 nm, the photoacoustic microscope still has enough sensitivity to image small blood vessels while providing high optical absorption contrast between melanin and hemoglobin. Label-free melanoma cells in flowing bovine blood are imaged in vitro, yielding measurements of both cell size and flow speed.

In Vivo Photoacoustic Microscopy of Human Cutaneous Microvasculature and a Nevus

Journal of Biomedical Optics. Jan-Feb, 2011  |  Pubmed ID: 21280921

In several human volunteers, photoacoustic microscopy (PAM) has been utilized for noninvasive cutaneous imaging of the skin microvasculature and a melanocytic nevus. Microvascular networks in both acral and nonacral skin were imaged, and multiple features within the skin have been identified, including the stratum corneum, epidermal-dermal junction, and subpapillary vascular plexus. Several vascular and structural differences between acral and nonacral skin were also observed in the photoacoustic images. In addition, a nevus was photoacoustically imaged, excised, and histologically analyzed. The photoacoustic images allowed for in vivo measurement of tumor thickness, depth, and microvasculature-values confirmed by histologic examination. The presented images demonstrate the potential of PAM to aid in the study and evaluation of cutaneous microcirculation and analysis of pigmented lesions. Through its ability to three-dimensionally image the structure and function of the microvasculature and pigmented lesions, PAM can have a clinical impact in diagnosis and assessment of systemic diseases that affect the microvasculature such as diabetes and cardiovascular disease, cutaneous malignancies such as melanoma, and potentially other skin disorders.

Conditional HIF-1 Induction Produces Multistage Neovascularization with Stage-specific Sensitivity to VEGFR Inhibitors and Myeloid Cell Independence

Blood. Apr, 2011  |  Pubmed ID: 21307392

Neovascularization is a crucial component of tumor growth and ischemia. Although prior work primarily used disease models, delineation of neovascularization in the absence of disease can reveal intrinsic mechanisms of microvessel regulation amenable to manipulation in illness. We created a conditional model of epithelial HIF-1 induction in adult mice (TetON-HIF-1 mice). Longitudinal photoacoustic microscopy (L-PAM) was coincidentally developed for noninvasive, label-free serial imaging of red blood cell-perfused vasculature in the same mouse for weeks to months. TetON-HIF-1 mice evidenced 3 stages of neovascularization: development, maintenance, and transgene-dependent regression. Regression occurred despite extensive and tight pericyte coverage. L-PAM mapped microvascular architecture and quantified volumetric changes in neocapillary morphogenesis, arteriovenous remodeling, and microvessel regression. Developmental stage endothelial proliferation down-regulation was associated with a DNA damage checkpoint consisting of p53, p21, and endothelial γ-H2AX induction. The neovasculature was temporally responsive to VEGFR2 immuno-blockade, with the developmental stage sensitive, and the maintenance stage resistant, to DC101 treatment. L-PAM analysis also pinpointed microvessels ablated or resistant to VEGFR2 immuno-blockade. HIF-1-recruited myeloid cells did not mediate VEGFR2 inhibitor resistance. Thus, HIF-1 neovascularization in the absence of disease is self-regulated via cell autonomous endothelial checkpoints, and resistant to angiogenesis inhibitors independent of myeloid cells.

In Vivo Functional Photoacoustic Microscopy of Cutaneous Microvasculature in Human Skin

Journal of Biomedical Optics. Feb, 2011  |  Pubmed ID: 21361688

Microcirculation is an important component of the cardiovascular system and can be used to assess systemic cardiovascular health. Numerous studies have investigated cutaneous microcirculation as an indicator of cardiovascular related diseases. Such research has shown promising results; however, there are many limitations regarding the employed measurement techniques, such as poor depth and spatial resolution and measurement versatility. Here we show the results of functional cutaneous microvascular experiments measured with photoacoustic microscopy, which provides high spatial resolution and multiparameter measurements. In a set of experiments, microvascular networks located in the palms of volunteers were perturbed by periodic ischemic events, and the subsequent hemodynamic response to the stimulus was recorded. Results indicate that during periods of arterial occlusion, the relative oxygen saturation of the capillary vessels decreased below resting levels, and temporarily increased above resting levels immediately following the occlusion. Furthermore, a hyperemic reaction to the occlusions was measured, and the observation agreed well with similar measurements using more conventional imaging techniques. Due to its exceptional capability to functionally image vascular networks with high spatial resolution, photoacoustic microscopy could be a beneficial biomedical tool to assess microvascular functioning and applied to patients with diseases that affect cardiovascular health. © 2011 Society of Photo-Optical Instrumentation Engineers.

Noninvasive Photoacoustic and Fluorescence Sentinel Lymph Node Identification Using Dye-loaded Perfluorocarbon Nanoparticles

ACS Nano. Jan, 2011  |  Pubmed ID: 21171567

The contrast mechanisms used for photoacoustic tomography (PAT) and fluorescence imaging differ in subtle, but significant, ways. The design of contrast agents for each or both modalities requires an understanding of the spectral characteristics as well as intra- and intermolecular interactions that occur during formulation. We found that fluorescence quenching that occurs in the formulation of near-infrared (NIR) fluorescent dyes in nanoparticles results in enhanced contrast for PAT. The ability of the new PAT method to utilize strongly absorbing chromophores for signal generation allowed us to convert a highly fluorescent dye into an exceptionally high PA contrast material. Spectroscopic characterization of the developed NIR dye-loaded perfluorocarbon-based nanoparticles for combined fluorescence and PA imaging revealed distinct dye-dependent photophysical behavior. We demonstrate that the enhanced contrast allows detection of regional lymph nodes of rats in vivo with time-domain optical and photoacoustic imaging methods. The results further show that the use of fluorescence lifetime imaging, which is less dependent on fluorescence intensity, provides a strategic approach to bridge the disparate contrast reporting mechanisms of fluorescence and PA imaging methods.

Gold Nanocages Covered with Thermally-responsive Polymers for Controlled Release by High-intensity Focused Ultrasound

Nanoscale. Apr, 2011  |  Pubmed ID: 21321760

This paper describes the use of Au nanocages covered with smart, thermally-responsive polymers for controlled release with high-intensity focused ultrasound (HIFU). HIFU is a highly precise medical procedure that uses focused ultrasound to heat and destroy pathogenic tissue rapidly and locally in a non-invasive or minimally invasive manner. The released dosage could be remotely controlled by manipulating the power of HIFU and/or the duration of exposure. We demonstrated localized release within the focal volume of HIFU by using gelatin phantom samples containing dye-loaded Au nanocages. By placing chicken breast tissues on top of the phantoms, we further demonstrated the feasibility of this system for controlled release at depths up to 30 mm. Because it can penetrate more deeply into soft tissues than near-infrared light, HIFU is a potentially more effective external stimulus for rapid, on-demand drug release.

In-vivo Characterization of Optical Properties of Pigmented Skin Lesions Including Melanoma Using Oblique Incidence Diffuse Reflectance Spectrometry

Journal of Biomedical Optics. Feb, 2011  |  Pubmed ID: 21361657

In this letter, we report the first use of oblique incidence diffuse reflectance spectrometry to conduct in-vivo measurements of optical properties of three different types of pigmented skin lesions, including melanoma, dysplastic, and common nevi. Both absorption and reduced scattering coefficient spectra were estimated from the spatially resolved diffuse reflectance within the wavelength range of 455-765 nm for 144 pigmented skin lesions including 16 melanomas. The absorption and reduced scattering spectra were found to change with the malignancy of the skin lesions, which were generally higher for the malignant cases than the benign ones. Based on the measurement results, the physiological origin leading to the change of the absorption and scattering properties is also discussed.

Single-wavelength Functional Photoacoustic Microscopy in Biological Tissue

Optics Letters. Mar, 2011  |  Pubmed ID: 21368977

Recently, we developed a reflection-mode relaxation photoacoustic microscope, based on saturation intensity, to measure picosecond relaxation times using a nanosecond laser. Here, using the different relaxation times of oxygenated and deoxygenated hemoglobin molecules, both possessing extremely low fluorescence quantum yields, the oxygen saturation was quantified in vivo with single-wavelength photoacoustic microscopy. All previous functional photoacoustic microscopy measurements required imaging with multiple-laser-wavelength measurements to quantify oxygen saturation. Eliminating the need for multiwavelength measurements removes the influence of spectral properties on oxygenation calculations and improves the portability and cost-effectiveness of functional or molecular photoacoustic microscopy.

A New Theranostic System Based on Gold Nanocages and Phase-change Materials with Unique Features for Photoacoustic Imaging and Controlled Release

Journal of the American Chemical Society. Apr, 2011  |  Pubmed ID: 21401092

This communication reports a new theranostic system with a combination of capabilities to both enhance the contrast of photoacoustic (PA) imaging and control the release of a chemical or biological effector by high-intensity focused ultrasound (HIFU). The fabrication of this system simply involves filling the hollow interiors of gold nanocages with a phase-change material (PCM) such as 1-tetradecanol that has a melting point of 38-39 °C. The PCM can be premixed and thus loaded with a dye, as well as other chemical or biological effectors. When exposed to direct heating or HIFU, the PCM will melt and escape from the interiors of nanocages through small pores on the surface, concurrently releasing the encapsulated molecules into the surrounding medium. We can control the release profile by varying the power of HIFU, the duration of exposure to HIFU, or both.

Porphysome Nanovesicles Generated by Porphyrin Bilayers for Use As Multimodal Biophotonic Contrast Agents

Nature Materials. Apr, 2011  |  Pubmed ID: 21423187

Optically active nanomaterials promise to advance a range of biophotonic techniques through nanoscale optical effects and integration of multiple imaging and therapeutic modalities. Here, we report the development of porphysomes; nanovesicles formed from self-assembled porphyrin bilayers that generated large, tunable extinction coefficients, structure-dependent fluorescence self-quenching and unique photothermal and photoacoustic properties. Porphysomes enabled the sensitive visualization of lymphatic systems using photoacoustic tomography. Near-infrared fluorescence generation could be restored on dissociation, creating opportunities for low-background fluorescence imaging. As a result of their organic nature, porphysomes were enzymatically biodegradable and induced minimal acute toxicity in mice with intravenous doses of 1,000 mg kg(-1). In a similar manner to liposomes, the large aqueous core of porphysomes could be passively or actively loaded. Following systemic administration, porphysomes accumulated in tumours of xenograft-bearing mice and laser irradiation induced photothermal tumour ablation. The optical properties and biocompatibility of porphysomes demonstrate the multimodal potential of organic nanoparticles for biophotonic imaging and therapy.

In Vivo Integrated Photoacoustic and Confocal Microscopy of Hemoglobin Oxygen Saturation and Oxygen Partial Pressure

Optics Letters. Apr, 2011  |  Pubmed ID: 21478972

We developed dual-modality microscope integrating photoacoustic microscopy (PAM) and fluorescence confocal microscopy (FCM) to noninvasively image hemoglobin oxygen saturation (sO₂) and oxygen partial pressure (pO₂) in vivo in single blood vessels with high spatial resolution. While PAM measures sO₂ by imaging hemoglobin optical absorption at two wavelengths, FCM quantifies pO₂ using phosphorescence quenching. The variations of sO₂ and pO₂ values in multiple orders of vessel branches under hyperoxic (100% oxygen) and normoxic (21% oxygen) conditions correlate well with the oxygen-hemoglobin dissociation curve. In addition, the total concentration of hemoglobin is imaged by PAM at an isosbestic wavelength.

Second-generation Optical-resolution Photoacoustic Microscopy with Improved Sensitivity and Speed

Optics Letters. Apr, 2011  |  Pubmed ID: 21479007

We developed second-generation (G2) optical-resolution photoacoustic microscopy (OR-PAM). Incorporation of a novel acoustic detection scheme improved upon the sensitivity of our first-generation (G1) system by 18.4 dB, deepening the in vivo tissue penetration to 1.2 mm at 570 nm. Moreover, translating the imaging head instead of the living object accelerated the scanning speed by a factor of 5, widening the field of view within the same acquisition time. Mouse ears, as well as mouse brains with intact craniums, were imaged in vivo in both total concentration and oxygen saturation of hemoglobin.

Real-time Four-dimensional Optical-resolution Photoacoustic Microscopy with Au Nanoparticle-assisted Subdiffraction-limit Resolution

Optics Letters. Apr, 2011  |  Pubmed ID: 21479008

Photoacoustic microscopy (PAM) offers label-free, optical absorption contrast. A high-speed, high-resolution PAM system in an inverted microscope configuration with a laser pulse repetition rate of 100,000 Hz and a stationary ultrasonic transducer was built. Four-dimensional in vivo imaging of microcirculation in mouse skin was achieved at 18 three-dimensional volumes per second with repeated two-dimensional (2D) raster scans of 100 by 50 points. The corresponding 2D B-scan (50 A-lines) frame rate was 1800 Hz, and the one-dimensional A-scan rate was 90,000 Hz. The lateral resolution is 0.23 ± 0.03 μm for Au nanowire imaging, which is 2.0 times below the diffraction limit.

Multifocal Optical-resolution Photoacoustic Microscopy in Vivo

Optics Letters. Apr, 2011  |  Pubmed ID: 21479041

Although ultrasound arrays have been exploited in photoacoustic imaging to improve imaging speed, ultrasound-array-based optical-resolution photoacoustic microscopy (OR-PAM) has never been achieved previously to our knowledge. Here we present our development of multifocal OR-PAM using a microlens array for optical illumination and an ultrasound array for photoacoustic detection. Our system is capable of imaging hemoglobin concentration and oxygenation in individual microvessels in vivo at high speed. Compared with a single focus, multiple foci reduce the scanning load and increase the imaging speed significantly. The current multifocal system can acquire 1000 × 500 × 200 voxels at ~10 μm lateral resolution within 4 min.

Photoacoustic Tomography of Foreign Bodies in Soft Biological Tissue

Journal of Biomedical Optics. Apr, 2011  |  Pubmed ID: 21529086

In detecting small foreign bodies in soft biological tissue, ultrasound imaging suffers from poor sensitivity (52.6%) and specificity (47.2%). Hence, alternative imaging methods are needed. Photoacoustic (PA) imaging takes advantage of strong optical absorption contrast and high ultrasonic resolution. A PA imaging system is employed to detect foreign bodies in biological tissues. To achieve deep penetration, we use near-infrared light ranging from 750 to 800 nm and a 5-MHz spherically focused ultrasonic transducer. PA images were obtained from various targets including glass, wood, cloth, plastic, and metal embedded more than 1 cm deep in chicken tissue. The locations and sizes of the targets from the PA images agreed well with those of the actual samples. Spectroscopic PA imaging was also performed on the objects. These results suggest that PA imaging can potentially be a useful intraoperative imaging tool to identify foreign bodies.

Time-reversed Ultrasonically Encoded Optical Focusing into Scattering Media

Nature Photonics. Mar, 2011  |  Pubmed ID: 21532925

Light focusing plays a central role in biomedical imaging, manipulation, and therapy. In scattering media, direct light focusing becomes infeasible beyond one transport mean free path. All previous methods1-3 to overcome this diffusion limit lack a practical internal "guide star."4 Here we proposed and experimentally validated a novel concept, called Time-Reversed Ultrasonically Encoded (TRUE) optical focusing, to deliver light into any dynamically defined location inside a scattering medium. First, diffused coherent light is encoded by a focused ultrasonic wave to provide a virtual internal "guide star"; then, only the encoded light is time-reversed and transmitted back to the ultrasonic focus. The TRUE optical focus-defined by the ultrasonic wave-is unaffected by multiple scattering of light. Such focusing is especially desirable in biological tissue where ultrasonic scattering is ~1000 times weaker than optical scattering. Various fields including biomedical and colloidal optics can benefit from TRUE optical focusing.

In Vivo Photoacoustic Mapping of Lymphatic Systems with Plasmon-resonant Nanostars

Journal of Materials Chemistry. Jan, 2011  |  Pubmed ID: 21660122

Plasmon-resonant nanostars (NSTs) provide excellent contrast enhancement for photoacoustic tomography. The high photoacoustic sensitivity of NSTs at near-infrared wavelengths enable their in vivo detection in rat sentinel lymph nodes and vessels, with direct application toward lymphangiography.

Noninvasive Photoacoustic Microscopy of Living Cells in Two and Three Dimensions Through Enhancement by a Metabolite Dye

Angewandte Chemie (International Ed. in English). Aug, 2011  |  Pubmed ID: 21681876

In Vivo Photoacoustic Tomography of Mouse Cerebral Edema Induced by Cold Injury

Journal of Biomedical Optics. Jun, 2011  |  Pubmed ID: 21721821

For the first time, we have implemented photoacoustic tomography (PAT) to image the water content of an edema in vivo. We produced and imaged a cold-induced cerebral edema transcranially, then obtained blood vessel and water accumulation images at 610 and 975 nm, respectively. We tracked the changes at 12, 24, and 36 h after the cold injury. The blood volume decreased after the cold injury, and the maximum area of edema was observed 24 h after the cold injury. We validated PAT of the water content of the edema through magnetic Resonance Imaging and the water spectrum from the spectrophotometric measurement.

Label-free Bond-selective Imaging by Listening to Vibrationally Excited Molecules

Physical Review Letters. Jun, 2011  |  Pubmed ID: 21770549

We report the realization of vibrational photoacoustic (VPA) microscopy using optical excitation of molecular overtone vibration and acoustic detection of the resultant pressure transients. Our approach eliminates the tissue scattering problem encountered in near-infrared spectroscopy and enables depth-resolved signal collection. The 2nd overtone of the CH bond stretch around 8300  cm(-1), where blood interference is minimal, is excited. We demonstrate 3D VPA imaging of lipid-rich atherosclerotic plaques by excitation from the artery lumen, and lipid storage in live Drosophila larvae, with millimeter scale penetration depth [corrected].

VEGF is Essential for Hypoxia-inducible Factor-mediated Neovascularization but Dispensable for Endothelial Sprouting

Proceedings of the National Academy of Sciences of the United States of America. Aug, 2011  |  Pubmed ID: 21784979

Although our understanding of the molecular regulation of adult neovascularization has advanced tremendously, vascular-targeted therapies for tissue ischemia remain suboptimal. The master regulatory transcription factors of the hypoxia-inducible factor (HIF) family are attractive therapeutic targets because they coordinately up-regulate multiple genes controlling neovascularization. Here, we used an inducible model of epithelial HIF-1 activation, the TetON-HIF-1 mouse, to test the requirement for VEGF in HIF-1 mediated neovascularization. TetON-HIF-1, K14-Cre, and VEGF(flox/flox) alleles were combined to create TetON-HIF-1:VEGF(Δ) mice to activate HIF-1 and its target genes in adult basal keratinocytes in the absence of concomitant VEGF. HIF-1 induction failed to produce neovascularization in TetON-HIF-1:VEGF(Δ) mice despite robust up-regulation of multiple proangiogenic HIF targets, including PlGF, adrenomedullin, angiogenin, and PAI-1. In contrast, endothelial sprouting was preserved, enhanced, and more persistent, consistent with marked reduction in Dll4-Notch-1 signaling. Optical-resolution photoacoustic microscopy, which provides noninvasive, label-free, high resolution, and wide-field vascular imaging, revealed the absence of both capillary expansion and arteriovenous remodeling in serially imaged individual TetON-HIF-1:VEGF(Δ) mice. Impaired TetON-HIF-1:VEGF(Δ) neovascularization could be partially rescued by 12-O-tetradecanoylphorbol-13-acetate skin treatment. These data suggest that therapeutic angiogenesis for ischemic cardiovascular disease may require treatment with both HIF-1 and VEGF.

Photoacoustic Microscopy of Microvascular Responses to Cortical Electrical Stimulation

Journal of Biomedical Optics. Jul, 2011  |  Pubmed ID: 21806263

Advances in the functional imaging of cortical hemodynamics have greatly facilitated the understanding of neurovascular coupling. In this study, label-free optical-resolution photoacoustic microscopy (OR-PAM) was used to monitor microvascular responses to direct electrical stimulations of the mouse somatosensory cortex through a cranial opening. The responses appeared in two forms: vasoconstriction and vasodilatation. The transition between these two forms of response was observed in single vessels by varying the stimulation intensity. Marked correlation was found between the current-dependent responses of two daughter vessels bifurcating from the same parent vessel. Statistical analysis of twenty-seven vessels from three different animals further characterized the spatial-temporal features and the current dependence of the microvascular response. Our results demonstrate that OR-PAM is a valuable tool to study neurovascular coupling at the microscopic level.

Label-free Oxygen-metabolic Photoacoustic Microscopy in Vivo

Journal of Biomedical Optics. Jul, 2011  |  Pubmed ID: 21806264

Almost all diseases, especially cancer and diabetes, manifest abnormal oxygen metabolism. Accurately measuring the metabolic rate of oxygen (MRO(2)) can be helpful for fundamental pathophysiological studies, and even early diagnosis and treatment of disease. Current techniques either lack high resolution or rely on exogenous contrast. Here, we propose label-free metabolic photoacoustic microscopy (mPAM) with small vessel resolution to noninvasively quantify MRO(2) in vivo in absolute units. mPAM is the unique modality for simultaneously imaging all five anatomical, chemical, and fluid-dynamic parameters required for such quantification: tissue volume, vessel cross-section, concentration of hemoglobin, oxygen saturation of hemoglobin, and blood flow speed. Hyperthermia, cryotherapy, melanoma, and glioblastoma were longitudinally imaged in vivo. Counterintuitively, increased MRO(2) does not necessarily cause hypoxia or increase oxygen extraction. In fact, early-stage cancer was found to be hyperoxic despite hypermetabolism.

Photoacoustic Tomography of Monkey Brain Using Virtual Point Ultrasonic Transducers

Journal of Biomedical Optics. Jul, 2011  |  Pubmed ID: 21806266

A photoacoustic tomography system (PAT) using virtual point ultrasonic transducers was developed and applied to image a monkey brain. The custom-built transducers provide a 10-fold greater field-of-view (FOV) than finite-aperture unfocused transducers as well as an improved signal-to-noise ratio (SNR) and reduced artifacts rather than negative-lens transducers. Their tangential resolution, radial resolution, and (SNR) improvements were quantified using tissue phantoms. Our PAT system can achieve high uniformity in both resolution (<1 mm) and SNR (>8) within a large FOV of 6 cm in diameter, even when the imaging objects are enclosed by a monkey skull. The cerebral cortex of a monkey brain was accurately mapped transcranially, through a skull ranging from 2 to 4 mm in thickness. This study demonstrates that PAT can overcome the optical and ultrasound attenuation of a relatively thick skull and can potentially be applied to human neonatal brain imaging.

Effects of Photoacoustic Imaging and Photothermal Ablation Therapy Mediated by Targeted Hollow Gold Nanospheres in an Orthotopic Mouse Xenograft Model of Glioma

Cancer Research. Oct, 2011  |  Pubmed ID: 21856744

Advancements in nanotechnology have made it possible to create multifunctional nanostructures that can be used simultaneously to image and treat cancers. For example, hollow gold nanospheres (HAuNS) have been shown to generate intense photoacoustic signals and induce efficient photothermal ablation (PTA) therapy. In this study, we used photoacoustic tomography, a hybrid imaging modality, to assess the intravenous delivery of HAuNS targeted to integrins that are overexpressed in both glioma and angiogenic blood vessels in a mouse model of glioma. Mice were then treated with near-infrared laser, which elevated tumor temperature by 20.7°C. We found that PTA treatment significantly prolonged the survival of tumor-bearing mice. Taken together, these results show the feasibility of using a single nanostructure for image-guided local tumor PTA therapy with photoacoustic molecular imaging.

Quantitative Photoacoustic Imaging: Correcting for Heterogeneous Light Fluence Distributions Using Diffuse Optical Tomography

Journal of Biomedical Optics. Sep, 2011  |  Pubmed ID: 21950930

The specificity of molecular and functional photoacoustic (PA) images depends on the accuracy of the photoacoustic absorption spectroscopy. The PA signal is proportional to the product of the optical absorption coefficient and local light fluence; quantitative PA measurements of the optical absorption coefficient therefore require an accurate estimation of optical fluence. Light-modeling aided by diffuse optical tomography (DOT) can be used to map the required fluence and to reduce errors in traditional PA spectroscopic analysis. As a proof-of-concept, we designed a tissue-mimicking phantom to demonstrate how fluence-related artifacts in PA images can lead to misrepresentations of tissue properties. To correct for these inaccuracies, the internal fluence in the tissue phantom was estimated by using DOT to reconstruct spatial distributions of the absorption and reduced scattering coefficients of multiple targets within the phantom. The derived fluence map, which only consisted of low spatial frequency components, was used to correct PA images of the phantom. Once calibrated to a known absorber, this method reduced errors in estimated absorption coefficients from 33% to 6%. These results experimentally demonstrate that combining DOT with PA imaging can significantly reduce fluence-related errors in PA images, while producing quantitatively accurate, high-resolution images of the optical absorption coefficient.

Performance Benchmarks of an Array-based Hand-held Photoacoustic Probe Adapted from a Clinical Ultrasound System for Non-invasive Sentinel Lymph Node Imaging

Philosophical Transactions. Series A, Mathematical, Physical, and Engineering Sciences. Nov, 2011  |  Pubmed ID: 22006911

Clinical translation of photoacoustic (PA) imaging can be facilitated by integration with commercial ultrasound (US) scanners to enable dual-modality imaging. An array-based US scanner was modified for hand-held PA imaging. The performance was benchmarked in terms of signal-to-noise ratio (SNR), axial spatial resolution and sensitivity. PA images of a tube, filled with methylene blue (MB; approx. 30 mM) and placed at various depths in chicken tissue, were acquired. A 5 cm penetration depth was achieved with an 18.6 dB SNR using a laser fluence of 3 mJ cm(-2), only one-seventh of the safety limit (20 mJ cm(-2)). An axial resolution of approximately 400 μm was maintained at all imaging depths. The PA sensitivity to MB placed 2.3 cm deep in chicken tissue was less than 100 μM. Further, after intradermal injection of MB (approx. 30 mM), a rat sentinel lymph node was clearly identified in vivo, beneath a 3.8 cm thick layer of chicken breast. The accumulated concentration of MB in the node was estimated to be approximately 7 mM. The noise-equivalent sensitivities (approx. 2 cm depth) were 17 and 85 μM, ex vivo and in vivo, respectively. These results support the use of this PA system for non-invasive mapping and image-guided needle biopsy of sentinel nodes in breast cancer patients.

Gold Nanocages As Contrast Agents for Photoacoustic Imaging

Contrast Media & Molecular Imaging. Sep-Oct, 2011  |  Pubmed ID: 22025337

Gold nanoparticles with tunable absorption and scattering properties have been developed as contrast agents for various optical imaging techniques. As a hybrid modality that combines the merits of both optical and ultrasonic imaging, photoacoustic (PA) imaging also benefits from the use of these nanoparticles to greatly enhance the contrast for visualization of structures and biomarkers in biological tissues. Gold nanocages characterized by hollow interiors, ultrathin and porous walls are of particular interest for in vivo PA imaging because of their compact sizes, bio-inertness and well-defined surface chemistry, as well as their strong and highly wavelength-tunable optical absorption in the near-infrared (NIR) optical window of soft tissues. This review discusses the application of gold nanocages as a new class of contrast agents for PA imaging in the context of cancer diagnosis.

Functional Photoacoustic Microscopy of PH

Journal of Biomedical Optics. Oct, 2011  |  Pubmed ID: 22029342

pH is a tightly regulated indicator of metabolic activity. In mammalian systems, an imbalance of pH regulation may result from or result in serious illness. In this paper, we report photoacoustic microscopy (PAM) of a commercially available pH-sensitive fluorescent dye (SNARF-5F carboxylic acid) in tissue phantoms. We demonstrated that PAM is capable of pH imaging in absolute values at tissue depths of up to 2.0 mm, greater than possible with other forms of optical microscopy.

In Vivo Quantitative Evaluation of the Transport Kinetics of Gold Nanocages in a Lymphatic System by Noninvasive Photoacoustic Tomography

ACS Nano. Dec, 2011  |  Pubmed ID: 22054348

Sentinel lymph node (SLN) biopsy has emerged as a preferred method for axillary lymph node staging of breast cancer, and imaging the SLN in three-dimensional space is a prerequisite for the biopsy. Conventional SLN mapping techniques based on the injection of an organic dye or a suspension of radioactive colloids suffer from invasive surgical operation for visual detection of the dye or hazardous radioactive components and low spatial resolution of Geiger counters in detecting the radioactive colloids. This work systematically investigates the use of gold nanocages (AuNCs) as a novel class of optical tracers for noninvasive SLN imaging by photoacoustic (PA) tomography in a rat model. The transport of AuNCs in a lymphatic system and uptake by the SLN were evaluated by PA tomography on the axillary region of a rat. Quantification of AuNCs accumulated in the lymph node was achieved by correlating the data from PA imaging with the results from inductively coupled plasma mass spectrometry. Several parameters were systematically evaluated and optimized, including the concentration, size, and surface charge of the AuNCs. These results are critical to the further development of this AuNC-based PA tomography system for noninvasive SLN imaging, providing valuable information for metastatic cancer staging.

Molecular Photoacoustic Imaging of Angiogenesis with Integrin-targeted Gold Nanobeacons

FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology. Mar, 2011  |  Pubmed ID: 21097518

Photoacoustic tomography (PAT) combines optical and acoustic imaging to generate high-resolution images of microvasculature. Inherent sensitivity to hemoglobin permits PAT to image blood vessels but precludes discriminating neovascular from maturing microvasculature. α(v)β(3)-Gold nanobeacons (α(v)β(3)-GNBs) for neovascular molecular PAT were developed, characterized, and demonstrated in vivo using a mouse Matrigel-plug model of angiogenesis. PAT results were microscopically corroborated with fluorescent α(v)β(3)-GNB localization and supporting immunohistology in Rag1(tm1Mom) Tg(Tie-2-lacZ)182-Sato mice. α(v)β(3)-GNBs (154 nm) had 10-fold greater contrast than blood on an equivolume basis when imaged at 740 nm to 810 nm in blood. The lowest detectable concentration in buffer was 290 nM at 780 nm. Noninvasive PAT of angiogenesis using a 10-MHz ultrasound receiver with α(v)β(3)-GNBs produced a 600% increase in signal in a Matrigel-plug mouse model relative to the inherent hemoglobin contrast pretreatment. In addition to increasing the contrast of neovessels detected at baseline, α(v)β(3)-GNBs allowed visualization of numerous angiogenic sprouts and bridges that were undetectable before contrast injection. Competitive inhibition of α(v)β(3)-GNBs with α(v)β(3)-NBs (no gold particles) almost completely blocked contrast enhancement to pretreatment levels, similar to the signal from animals receiving saline only. Consistent with other studies, nontargeted GNBs passively accumulated in the tortuous neovascular but provided less than half of the contrast enhancement of the targeted agent. Microscopic studies revealed that the vascular constrained, rhodamine-labeled α(v)β(3)-GNBs homed specifically to immature neovasculature (PECAM(+), Tie-2(-)) along the immediate tumor periphery, but not to nearby mature microvasculature (PECAM(+), Tie-2(+)). The combination of PAT and α(v)β(3)-GNBs offered sensitive and specific discrimination and quantification of angiogenesis in vivo, which may be clinically applicable to a variety of highly prevalent diseases, including cancer and cardiovascular disease.

Three-dimensional Photoacoustic Tomography Based on the Focal-line Concept

Journal of Biomedical Optics. Sep, 2011  |  Pubmed ID: 21950908

A full ring ultrasonic array-based photoacoustic tomography system was recently developed for small animal brain imaging. The 512-element array is cylindrically focused in the elevational direction, and can acquire a two-dimensional (2D) image in 1.6 s. In this letter, we demonstrate the three-dimensional (3D) imaging capability of this system. A novel 3D reconstruction algorithm was developed based on the focal-line concept. Compared to 3D images acquired simply by stacking a series of 2D images, the 3D focal-line reconstruction method renders images with much less artifacts, and improves the elevational resolution by 30% and the signal-to-noise ratio by two times. The effectiveness of the proposed algorithm was first validated by numerical simulations and then demonstrated with a hair phantom experiment and an ex vivo mouse embryo experiment.

Photoacoustic Tomography: Fundamentals, Advances and Prospects

Contrast Media & Molecular Imaging. Sep-Oct, 2011  |  Pubmed ID: 22025335

Optical microscopy has been contributing to the development of life science for more than three centuries. However, due to strong optical scattering in tissue, its in vivo imaging ability has been restricted to studies at superficial depths. Advances in photoacoustic tomography (PAT) now allow multiscale imaging at depths from sub-millimeter to several centimeters, with spatial resolutions from sub-micrometer to sub-millimeter. Because of this high scalability and its unique optical absorption contrast, PAT is capable of performing anatomical, functional, molecular and fluid-dynamic imaging at various system levels, and is playing an increasingly important role in fundamental biological research and clinical practice. This review discusses recent technical progress in PAT and presents corresponding applications. It ends with a discussion of several prospects and their technical challenges.

Recent Advances in Colloidal Gold Nanobeacons for Molecular Photoacoustic Imaging

Contrast Media & Molecular Imaging. Sep-Oct, 2011  |  Pubmed ID: 22025338

Photoacoustic imaging (PAI) represents a hybrid, nonionizing modality, which has been of particular interest because of its satisfactory spatial resolution and high soft tissue contrast. PAI has the potential to provide both functional and molecular imaging in vivo since optical absorption is sensitive to physiological parameters. In this review we summarize our effort to advance molecular PAI with colloidal gold nanobeacons (GNB). GNB represents a robust nanoparticle platform that entraps multiple copies of tiny gold nanoparticles (2-4 nm) within a larger colloidal particle encapsulated by biocompatible synthetic or natural amphilines. The utilization of numerous small gold particles greatly amplifies the signal without exceeding the renal elimination threshold size. With fibrin-targeted GNB, the robust detection of microthrombus formed over a ruptured atherosclerotic plaque has been achieved, which offers an important opportunity to recognize patients with moderate lumen stenosis but high risk of stroke. With the use of second-generation smaller GNBs, the potential to improve sentinel lymph node assessment and biopsy was advanced with respect to rapidity and sensitivity of detection in mice. Finally, for angiogenesis, an essential microanatomical biomarker of tumor and cardiovascular disease progression, integrin-targeted GNBs allowed visualization of numerous angiogenic sprouts and bridges that were otherwise undetectable from inherent blood signal alone, offering sensitive and specific discrimination and quantification of angiogenesis in vivo.

Multiscale Photoacoustic Microscopy of Single-Walled Carbon Nanotube-Incorporated Tissue Engineering Scaffolds

Tissue Engineering. Part C, Methods. Dec, 2011  |  Pubmed ID: 22082018

Three-dimensional polymeric scaffolds provide structural support and function as substrates for cells and bioactive molecules necessary for tissue regeneration. Noninvasive real-time imaging of scaffolds and/or the process of tissue formation within the scaffold remains a challenge. Microcomputed tomography, the widely used technique to characterize polymeric scaffolds, shows poor contrast for scaffolds immersed in biological fluids, thereby limiting its utilities under physiological conditions. In this article, multiscale photoacoustic microscopy (PAM), consisting of both acoustic-resolution PAM (AR-PAM) and optical-resolution PAM (OR-PAM), was employed to image and characterize single-walled carbon-nanotube (SWNT)-incorporated poly(lactic-co-glycolic acid) polymer scaffolds immersed in biological buffer. SWNTs were incorporated to reinforce the mechanical properties of the scaffolds, and to enhance the photoacoustic signal from the scaffolds. By choosing excitation wavelengths of 570 and 638 nm, multiscale PAM could spectroscopically differentiate the photoacoustic signals generated from blood and from carbon-nanotube-incorporated scaffolds. OR-PAM, providing a fine lateral resolution of 2.6 μm with an adequate tissue penetration of 660 μm, successfully quantified the average porosity and pore size of the scaffolds to be 86.5%±1.2% and 153±15 μm in diameter, respectively. AR-PAM further extended the tissue penetration to 2 mm at the expense of lateral resolution (45 μm). Our results suggest that PAM is a promising tool for noninvasive real-time imaging and monitoring of tissue engineering scaffolds in vitro, and in vivo under physiological conditions.

Gold Nanostructures: a Class of Multifunctional Materials for Biomedical Applications

Chemical Society Reviews. Jan, 2011  |  Pubmed ID: 20818451

Gold nanostructures have proven to be a versatile platform for a broad range of biomedical applications, with potential use in numerous areas including: diagnostics and sensing, in vitro and in vivo imaging, and therapeutic techniques. These applications are possible because of the highly favorable properties of gold nanostructures, many of which can be tailored for specific applications. In the first part of this tutorial review, we will discuss the most critical properties of gold nanostructures for biomedical applications: surface chemistry, localized surface plasmon resonance (LSPR), and morphology. In the second part of the review, we will discuss how these properties can be harnessed for a selection of biomedical applications, aiming to give the reader an overview of general strategies as well as highlight some recent advances in this field.

Photoacoustic Microscopy of Tyrosinase Reporter Gene in Vivo

Journal of Biomedical Optics. Aug, 2011  |  Pubmed ID: 21895303

Photoacoustic tomography is a hybrid modality based on optical absorption excitation and ultrasonic detection. It is sensitive to melanin, one of the primary absorbers in skin. For cells that do not naturally contain melanin, melanin production can be induced by introducing the gene for tyrosinase, the primary enzyme responsible for expression of melanin in melanogenic cells. Optical resolution photoacoustic microscopy was used in the ex vivo study reported here, where the signal from transfected cells increased by more than 10 times over wild-type cells. A subsequent in vivo experiment was conducted to demonstrate the capability of photoacoustic microscopy to spectrally differentiate between tyrosinase-catalyzed melanin and various other absorbers in tissue.

Reflection-mode Time-reversed Ultrasonically Encoded Optical Focusing into Turbid Media

Journal of Biomedical Optics. Aug, 2011  |  Pubmed ID: 21895305

Time-reversed ultrasonically encoded (TRUE) optical focusing was recently proposed to deliver light dynamically to a tight region inside a scattering medium. In this letter, we report the first development of a reflection-mode TRUE optical focusing system. A high numerical aperture light guide is used to transmit the diffusely reflected light from a turbid medium to a phase-conjugate mirror (PCM), which is sensitive only to the ultrasound-tagged light. From the PCM, a phase conjugated wavefront of the tagged light is generated and conveyed by the same light guide back to the turbid medium, subsequently converging to the ultrasonic focal zone. We present experimental results from this system, which has the ability to focus light in a highly scattering medium with a round-trip optical penetration thickness (extinction coefficient multiplied by round-trip depth) as large as 160.

Time-reversed Ultrasonically Encoded Optical Focusing into Tissue-mimicking Media with Thickness Up to 70 Mean Free Paths

Journal of Biomedical Optics. Aug, 2011  |  Pubmed ID: 21895321

In turbid media such as biological tissue, multiple scattering hinders direct light focusing at depths beyond one transport mean free path. As a solution to this problem, time-reversed ultrasonically encoded (TRUE) optical focusing is proposed based on ultrasonic encoding of diffused laser light and optical time reversal. In TRUE focusing, a laser beam of long coherence length illuminates a turbid medium, where the incident light undergoes multiple scattering and part of it gets ultrasonically encoded within the ultrasonic focal zone. A conjugated wavefront of the ultrasonically encoded light is then generated by a phase conjugate mirror outside the medium, which traces back the trajectories of the ultrasonically encoded diffused light and converges light to the ultrasonic focal zone. Here, we report the latest experimental improvement in TRUE optical focusing that increases its penetration in tissue-mimicking media from a thickness of 3.75 to 7.00 mm. We also demonstrate that the TRUE focus depends on the focal diameter of the ultrasonic transducer.

Nonionizing Photoacoustic Cystography in Vivo

Optics Letters. Sep, 2011  |  Pubmed ID: 21931403

We demonstrate the feasibility of a novel and nonionizing process for bladder imaging in vivo, called photoacoustic cystography (PAC). Using a photoacoustic imaging system, we have successfully imaged a rat bladder filled with clinically used Methylene Blue (MB) dye. An image contrast of ~8 was achieved. Further, spectroscopic PAC confirmed the accumulation of MB in the bladder. Using a laser pulse energy of less than 1 mJ/cm² (1/20 of the ANSI safety limit), a deeply (1.2 cm) positioned bladder in biological tissues was clearly visible in the PA image. Our results suggest that PAC can potentially provide a nonionizing, relatively cheap, and portable tool for bladder mapping. Among our clinical interests, nonionizing PAC with an injection of MB can potentially monitor vesicoureteral reflux in children.

In Vivo Imaging of Epileptic Activity Using 2-NBDG, a Fluorescent Deoxyglucose Analog

Journal of Neuroscience Methods. Jan, 2012  |  Pubmed ID: 21939688

Accurately locating epileptic foci has great importance in advancing the treatment of epilepsy. In this study, epileptic seizures were first induced by intracortical injection of 4-aminopyridine in rats. A fluorescent deoxyglucose substitute, 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (2-NBDG), was then continuously injected via the tail vein. Brain glucose metabolism was subsequently monitored by fluorescence imaging of 2-NBDG. The initial uptake rate of 2-NBDG at the injection site of 4-aminopyridine significantly exceeded that of the control injection site, which indicated local hypermetabolism induced by seizures. Our results show that 2-NBDG can be used for localizing epileptic foci.

Deep-tissue Photoacoustic Tomography of a Genetically Encoded Near-infrared Fluorescent Probe

Angewandte Chemie (International Ed. in English). Feb, 2012  |  Pubmed ID: 22213541

Skin-deep: The combination of a near-infrared fluorescent protein (iRFP) and deep-tissue photoacoustic tomography clearly demonstrates the superiority of iRFP over other genetically encoded probes. Impressive resolution (280 μm lateral and 75 μm axial) was obtained at a depth of 4 mm in a live animal, and volumetric images of a tumor were produced, thus allowing the spatially resolved monitoring of its development.

Photoacoustic Sentinel Lymph Node Imaging with Self-Assembled Copper Neodecanoate Nanoparticles

ACS Nano. Jan, 2012  |  Pubmed ID: 22229462

Photoacoustic tomography (PAT) is emerging as a novel, hybrid, and non-ionizing imaging modality because of its satisfactory spatial resolution and high soft tissue contrast. PAT combines the advantages of both optical and ultrasonic imaging methods. It opens up the possibilities for noninvasive staging of breast cancer and may replace sentinel lymph node (SLN) biopsy in clinic in the near future. In this work, we demonstrate for the first time that copper can be used as a contrast metal for near-infrared detection of SLN using PAT. A unique strategy is adopted to encapsulate multiple copies of Cu as organically soluble small molecule complexes within a phospholipid-entrapped nanoparticle. The nanoparticles assumed a size of 80-90 nm, which is the optimum hydrodynamic diameter for its distribution throughout the lymphatic systems. These particles provided at least 6-fold higher signal sensitivity in comparison to blood, which is a natural absorber of light. We also demonstrated that high SLN detection sensitivity with PAT can be achieved in a rodent model. This work clearly demonstrates for the first time the potential use of copper as an optical contrast agent.