Showing papers by "Stephen A. Boppart published in 2004"

Optical coherence tomography: feasibility for basic research and image-guided surgery of breast cancer.

Abstract: Diagnostic trends in medicine are being directed toward cellular and molecular processes, where treatment regimens are more amenable for cure. Optical imaging is capable of performing cellular and molecular imaging using the short wavelengths and spectroscopic properties of light. Diffuse optical tomography is an optical imaging technique that has been pursued as an alternative to X-ray mammography. While this technique permits non-invasive optical imaging of the whole breast, to date it is incapable of resolving features at the cellular level. Optical coherence tomography (OCT) is an emerging high-resolution biomedical imaging technology that for larger and undifferentiated cells can perform cellular-level imaging at the expense of imaging depth. OCT performs optical ranging in tissue and is analogous to ultrasound except reflections of near-infrared light are detected rather than sound. In this paper, an overview of the OCT technology is provided, followed by images demonstrating the feasibility of using OCT to image cellular features indicative of breast cancer. OCT images of a well-established carcinogen-induced rat mammary tumor model were acquired. Images from this common experimental model show strong correlation with corresponding histopathology. These results illustrate the potential of OCT for a wide range of basic research studies and for intra-operative image-guidance to identify foci of tumor cells within surgical margins during the surgical treatment of breast cancer.

Nonlinear interferometric vibrational imaging.

Abstract: Coherent anti-Stokes Raman scattering (CARS) processes are "coherent," but the phase of the anti-Stokes radiation is lost by most incoherent spectroscopic CARS measurements. We propose a Raman microscopy imaging method called nonlinear interferometric vibrational imaging, which measures Raman spectra by obtaining the temporal anti-Stokes signal through nonlinear interferometry. With a more complete knowledge of the anti-Stokes signal, we show through simulations that a high-resolution Raman spectrum can be obtained of a molecule in a single pulse using broad band radiation. This could be useful for identifying the three-dimensional spatial distribution of molecular species in tissue.

Near-infrared dyes as contrast-enhancing agents for spectroscopic optical coherence tomography

Abstract: Optical coherence tomography (OCT) images of biological tissues often have low contrast. Spectroscopic optical coherence tomography (SOCT) methods have been developed to enhance contrast but remain limited because most tissues are not spectrally active in the frequency bands of laser sources commonly used in OCT. Near-infrared (NIR) dyes with absorption spectra features within the OCT source spectrum can be used for enhancing contrast in this situation. We introduce and demonstrate the use of NIR dyes as contrast agents for SOCT. Contrast-enhanced images are compared with fluorescence microscopy, demonstrating a link between SOCT and fluorescence imaging.

Nonlinear optical contrast enhancement for optical coherence tomography.

Abstract: We present a new interferometric technique for measuring Coherent Anti-Stokes Raman Scattering (CARS) and Second Harmonic Generation (SHG) signals. Heterodyne detection is employed to increase the sensitivity in both CARS and SHG signal detection, which can also be extended to different coherent processes. The exploitation of the mentioned optical nonlinearities for molecular contrast enhancement in Optical Coherence Tomography (OCT) is presented.

Multi-functional plasmon-resonant contrast agents for optical coherence tomography

Abstract: A method of forming an image of a sample, comprising: forming an image of a mixture, by exposing the mixture to electromagnetic radiation; wherein the mixture comprises the sample and plasmon-resonant nanoparticles, and wherein the electromagnetic radiation is in the frequency range of infra-red to ultraviolet light.

Structural and Functional Optical Imaging of Three-Dimensional Engineered Tissue Development

Abstract: A significant amount of the data collected by cell biologists and tissue engineers relies on invasive imaging techniques to visualize dynamic structural and functional properties in engineered tissues. We report the use of optical coherence tomography and the comparative use of confocal microscopy to nondestructively and noninvasively monitor the structural and functional characteristics of threedimensional engineered tissues over time. The engineered tissue model is composed of chitosan scaffolds and fibroblasts transfected with vinculin fused to green fluorescent protein. We image the developmental process of engineered tissues from changes of tissue microarchitecture to cell–matrix adhesions in three dimensions. These findings demonstrate the potential for optical coherence tomography in applications in cell and tissue biology, tissue engineering, and drug discovery.

Separation of absorption and scattering profiles in spectroscopic optical coherence tomography using a least-squares algorithm

Abstract: In spectroscopic optical coherence tomography, it is important and useful to separately estimate the absorption and the scattering properties of tissue. In this paper, we propose a least-squares fitting algorithm to separate absorption and scattering profiles when near-infrared absorbing dyes are used. The algorithm utilizes the broadband Ti:sapphire laser spectrum together with joint time-frequency analysis. Noise contribution to the final estimation was analyzed using simulation. The validity of our algorithm was demonstrated using both single-layer and multi-layer tissue phantoms.

Interferometric differentiation between resonant coherent anti-Stokes Raman scattering and nonresonant four-wave-mixing processes

Abstract: A major impediment to the use of coherent anti-Stokes Raman scattering (CARS) to identify biological molecules is that the illumination levels required to produce a measurable signal often also produce significant nonresonant background from the medium, especially from water. We present a method of nonlinear interferometry to differentiate between which components of the anti-Stokes signal are resonant and nonresonant. The technique takes advantage of the persistence of intermediate states involved in the resonant process. This method is applicable to most existing pulsed CARS illumination methods and provides for identification of resonant CARS. We demonstrate the method by examining the signals produced by acetone, which exhibits resonance, and water, which does not.

Structural and functional imaging of 3D microfluidic mixers using optical coherence tomography.

Abstract: To achieve high mixing efficiency in microfluidic devices, complex designs are often required. Microfluidic devices have been evaluated with light and confocal microscopy, but fluid-flow characteristics at different depths are difficult to separate from the en face images produced. By using optical coherence tomography (OCT), an imaging modality capable of imaging 3D microstructures at micrometer-scale resolutions over millimeter-size scales, we obtained 3D dynamic functional and structural data for three representative microfluidic mixers: a Y channel mixer, a 3D serpentine mixer, and a vortex mixer. In the serpentine mixer, OCT image analysis revealed that the mixing efficiency was linearly dependent on the Reynolds number, whereas it appeared to have exponential dependence when imaged with light microscopy. The visual overlap of fluid flows in light-microscopy images leads to an overestimation of the mixing efficiency, an effect that was eliminated with OCT imaging. Doppler OCT measurements determined velocity profiles at various points in the serpentine mixer. Mixing patterns in the vortex mixer were compared with light-microscopy and OCT image analysis. These results demonstrate that OCT can significantly improve the characterization of 3D microfluidic device structure and function.

Real-time digital signal processing-based optical coherence tomography and Doppler optical coherence tomography

Abstract: We present the development and use of a real-time digital signal processing (DSP)-based optical coherence tomography (OCT) and Doppler OCT system. Images of microstructure and transient fluid-flow profiles are acquired using the DSP architecture for real-time processing of computationally intensive calculations. This acquisition system is readily configurable for a wide range of real-time signal processing and image processing applications in OCT.

Adaptive spectral apodization for sidelobe reduction in optical coherence tomography images

Abstract: Often in imaging systems, the bandpass of the system is not uniform. In temporal coherence imaging methods such as optical coherence tomography, one would like to achieve the most spatially confined impulse response possible with a given source spectrum, minimizing sidelobes that blur adjacent features together. Typically the spectrum of the source is controlled in order to remove sidelobes from the measured interferogram. However, the measured interferogram is not necessarily the best estimate of the scattering density of the object. In this work, a sidelobe supression method is proposed and demonstrated to achieve low sidelobes even with highly nonuniform, non-Gaussian spectra.

Magnetic contrast agents for optical coherence tomography

Abstract: The magneto-mechanical effect is exploited as a means of producing background-free contrast in optical coherence tomography (OCT). Contrast agents consisting of iron-oxide particles and protein microspheres encapsulating colloidal iron-oxide have a sufficiently high magnetic susceptibility to be detected by modulation of a magnetic field gradient using a small solenoid coil. The externally-applied magnetic field mechanically rotates or translates these highly scattering contrast agents within the sample at the modulation frequency, which is subsequently detected as amplitude modulation of the OCT signal. Pairs of sequential axial scans (A-lines) are acquired with the magnetic field on and off, allowing one to build up a pair of images corresponding to the "on" and "off" states of the magnetic field. These image pairs are differenced to look for magnetic-specific effects, allowing one to distinguish the magnetic contrast agents from non-magnetic structures within the sample with a signal-to-background ratio of ~23dB. This technique has the potential to be very powerful when coupled with targeting for in vivo molecular imaging. To evaluate this potential we demonstrate in vitro imaging of magnetically-labeled macrophage cells embedded in a 3D tissue phantom, in vitro tissue doped with contrast agents, and in vivo imaging of Xenopus laevis (African frog) tadpoles.

Retinal response of Macaca mulatta to picosecond laser pulses of varying energy and spot size

Abstract: We investigate the relationship between the laser beam at the retina (spot size) and the extent of retinal injury from single ul- trashort laser pulses. From previous studies it is believed that the reti- nal effect of single 3-ps laser pulses should vary in extent and loca- tion, depending on the occurrence of laser-induced breakdown (LIB) at the site of laser delivery. Single 3-ps pulses of 580-nm laser energy are delivered over a range of spot sizes to the retina of Macaca mu- latta. The retinal response is captured sequentially with optical coher- ence tomography (OCT). The in vivo OCT images and the extent of pathology on final microscopic sections of the laser site are com- pared. With delivery of a laser pulse with peak irradiance greater than that required for LIB, OCT and light micrographs demonstrate inner retinal injury with many intraretinal and/or vitreous hemorrhages. In contrast, broad outer retinal injury with minimal to no choriocapillaris effect is seen after delivery of laser pulses to a larger retinal area (60 to 300 mm diam) when peak irradiance is less than that required for LIB. The broader lesions extend into the inner retina when higher energy delivery produces intraretinal injury. Microscopic examination of stained fixed tissues provide better resolution of retinal morphology than OCT. OCT provides less resolution but could be guided over an in vivo, visible retinal lesion for repeated sampling over time during the evolution of the lesion formation. For 3-ps visible wavelength laser pulses, varying the spot size and laser energy directly affects the ex- tent of retinal injury. This again is believed to be partly due to the onset of LIB, as seen in previous studies. Spot-size dependence should be considered when comparing studies of retinal effects or when pur- suing a specific retinal effect from ultrashort laser pulses. © 2004 Society

Comparative performance analysis of time-frequency distributions for spectroscopic optical coherence tomography

Abstract: Joint time-frequency distributions (TFDs) can optimize the tradeoff between spectral and spatial resolution in spectroscopic OCT. Performances by representative TFDs are compared and optimized for different spectroscopic OCT applications.

Pulse shaping strategies for nonlinear interferometric vibrational imaging optimized for biomolecular imaging

Abstract: Nonlinear interferometric vibrational imaging (NIVI) measures the temporal cross-correlation of anti-Stokes radiation from coherent anti-Stokes Raman scattering (CARS) processes to achieve increased sensitivity, stray light rejection, and nonresonant background rejection. Because the intensity of CARS radiation is proportional to the square of the molecular density of a target resonance, it is critical to maximize the recoverable signal for a given illumination level. Especially if one desires to measure several resonances, there can be a sensitivity as well as a speed advantage to measuring them simultaneously rather than serially. We discuss the methods of sample excitation that NIVI allows and their potential sensitivity advantages, as well as present experimental results demonstrating Raman signal recovery using these pulse sequences.

Nonlinear interferometric vibrational imaging of molecular species

Abstract: Vibrationally-sensitive spectroscopic techniques are becoming important clinical tools for real-time, in vivo diagnostics. The molecular information made available with these techniques can provide early diagnostic signs of disease, often before morphological changes occur. We model and experimentally demonstrate a new technique for measuring optical spectroscopy signals using interferometric ranging. This new technique, nonlinear interferometric vibrational imaging (NIVI), uses principles from coherent anti-Stokes Raman scattering (CARS) spectroscopy and optical coherence tomography (OCT) to achieve cross-sectional imaging of the distribution of specific molecular species within a sample. Two CARS signals are generated, one from a known reference molecular species and a second from the unknown molecules in a sample. These coherent signals are interfered with each other using an interferometer setup. The intensity envelope of the interference signal provides a measure of the concentration of selected bonds present in the sample focal volume. The interference fringes themselves can provide phase information that will allow for the exact reconstruction of the vibrational characteristics of the molecules in the sample focal volume. Theoretical background to CARS interferometry is presented, the experimental laser systems are described, and a depth-resolved scan line of a benzene filled cuvette is demonstrated. The experimental results show close resemblance to the theoretical models. The advantages of NIVI over existing vibrational imaging systems and its clinical implications are discussed.

Interferometric contrast between resonant coherent anti-Stokes Raman sattering and nonresonant four-wave mixing

Abstract: Utilizing interferometry, we distinguish between resonant CARS and nonresonant four wave mixing, a pernicious problem of CARS microscopy. An optical parametric amplifier creates a coherent reference beam to demodulate the temporal CARS signal from acetone, sensitive to its concentration.

Contrast enhancement methods for optical coherence tomography

Abstract: Contrast in optical coherence tomography (OCT) images is often limited, particularly when pathological tissue is morphologically or optically similar to normal tissue. We present novel contrast enhancing methods designed to selectively identify tissues of interest.

Use of near-infrared fluorescent dyes in depth resolved spectroscopic optical coherence tomography

Abstract: Optical coherence tomography (OCT) images of biological tissues often have low contrast. Spectroscopic OCT (SOCT) methods have been developed to enhance contrast. Due to the time-frequency "uncertainty principle" in SOCT signal analysis, minute spectral changes can not be detected for high spatial resolution applications. In order to enhance SOCT contrast in tissues with low inherent spectral absorption, we developed a contrast enhancing technique using near-infrared (NIR) dyes. We developed an optimized algorithm for SOCT and an optimum dye concentration for imaging. The effect of contrast enhancement was studied both in tissue phantoms and in situ experiments. Contrast-enhanced images are compared with fluorescence microscopy, demonstrating a link between SOCT and fluorescence imaging.

Functional optical coherence tomography of stimulated and spontaneous scattering changes in neural tissue

Abstract: We demonstrate the use of functional optical coherence tomography (fOCT) for observing action potential propagation by detecting scattering changes in neural tissue. FOCT images of nerve fibers from the abdominal ganglion of the sea slug Aplysia californica were obtained before, during, and after electrical stimulation with monophasic as well as biphasic voltage pulses. A reversible localized increase in optical scattering was noted in the images obtained during stimulation compared to the images obtained before stimulation. In addition, M (motion)-mode images showed transient optical changes due to spontaneous electrical activity. To exclude local laser-induced temperature changes as a source for stimulation, we monitored the temperature effects of prolonged laser exposure with a thermistor and found that there was no substantial temperature increase. We conclude that OCT is sensitive to the optical changes induced in electrically stimulated nerve fibers, and that there is minimal tissue heating and no detectable damage caused by exposure to the laser.

A method for dynamically suppressing sidelobes in optical coherence tomography

Abstract: Resolution in optical coherence tomography is often degraded due to sidelobes of the point response. Frequently, the spectrum of the low-coherence source is unable to be changed to reduce sidelobes. We present a method that derives a space-invariant linear post processing digital filter that reduces the sidelobes in the reconstructed image while minimizing the increase in image noise. This method is demonstrated on the image of rat mammary tissue.

Imaging of mixing dynamics in micromixers using microscopy and optical coherence tomography

Abstract: We present the design of two efficient micromixers, a 3-D serpentine micromixer and a vortex mixer. Light and confocal microscopy and image analysis programs were used to study mixing efficiency in these two micromixers and a Y-shape straight channel. By utilizing Optical Coherence Tomography (OCT), an emerging high-resolution medical and biological imaging technology, we obtained 3-D structural data and mixing dynamics for the 3-D serpentine mixer and the vortex mixer. The results indicate that mixing efficiency characterized with OCT is more accurate.

Nonlinear optical contrast enhancement in OCT

Abstract: We report on a new interferometric technique for measuring Coherent Anti-Stokes Raman Scattering and Second Harmonic Generation signals. The exploitation of these optical nonlinearities for molecular contrast enhancement in Optical Coherence Tomography is presented.

In vivo detection of exogenous contrast agents using optical coherence tomography

Abstract: Contrast enhancement for in vivo OCT is investigated using plasmon-resonant gold nanorods, protein microspheres, liposomes, and iron-oxide particles. The spectroscopic, magnetomechanical and scattering properties are explored using in vivo rat, mouse and tadpole models.

Contrast enhancement and artifact reduction for projected index computed tomography

Abstract: The projected index computed tomography (PICT) imaging modality provides a computationally reconstructed image of optical index of refraction that is free from spatial distortions. Current implementation using standard filtered backprojection techniques contain linear artifacts resulting from beam path refraction at boundaries with a significant index change. We present the implementation of a least-squares solution whereby every projection datum is assigned a confidence weight, affecting the magnitude of its inclusion into the reconstructed image. The algorithm is shown to significantly reduce artifacts, hence increasing system sensitivity while maintaining spatial accuracy.

Multi-modality three-dimensional optical imaging of engineered tissues

Abstract: Optical coherence tomography (OCT), confocal, and multiphoton microscopy are used to nondestructively monitor in situ cell distribution and cell-matrix interaction in 3-D engineered tissues composed of chitosan scaffolds and fibroblasts transfected with GFP-tagged vinculin plasmid.

Functional optical coherence tomography of neurophysiology

Abstract: Functional optical coherence tomography (fOCT) is used to record optical changes that correspond to spontaneous and stimulated electrical activity in neural tissue. FOCT provides a high-resolution, real-time, non-invasive method for investigating neurophysiology.

Optical coherence tomography of breast cancer: feasibility for surgical guidance

Abstract: We present the use of OCT for breast cancer imaging. In situ rat mammary and excised human breast tumors were imaged. The feasibility of OCT for applications in surgical oncology of the breast is discussed.