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Because of its non-ionizing and molecular sensing nature, light has been an attractive tool in biomedicine. Scanning an optical focus allows not only high-resolution imaging but also manipulation and therapy. However, due to multiple photon scattering events, conventional optical focusing using an ordinary lens is limited to shallow depths of one transport mean free path (lt'), which corresponds to approximately 1 mm in human tissue. To overcome this limitation, ultrasonic modulation (or encoding) of diffuse light inside scattering media has enabled us to develop both deep-tissue optical imaging and focusing techniques, namely, ultrasound-modulated optical tomography (UOT) and time-reversed ultrasonically encoded (TRUE) optical focusing. While UOT measures the power of the encoded light to obtain an image, TRUE focusing generates a time-reversed (or phase-conjugated) copy of the encoded light, using a phase-conjugate mirror to focus light inside scattering media beyond 1 lt'. However, despite extensive progress in both UOT and TRUE focusing, the low signal-to-noise ratio in encoded-light detection remains a challenge to meeting both the speed and depth requirements for in vivo applications. This dissertation describes technological advancements of both UOT and TRUE focusing, in terms of their signal detection sensitivities, operational depths, and operational speeds. The first part of this dissertation describes sensitivity improvements of encoded-light detection in UOT, achieved by using a large area (~5 cm x 5 cm) photorefractive polymer. The photorefractive polymer allowed us to improve the detection etendue by more than 10 times that of previous detection schemes. It has enabled us to resolve absorbing objects embedded inside diffused media thicker than 80 lt', using moderate light power and short ultrasound pulses. The second part of this dissertation describes energy enhancement and fluorescent excitation using TRUE focusing in turbid media, using photorefractive materials as the phase-conjugate mirrors. By using a large-area photorefractive polymer as the phase-conjugate mirror, we boosted the focused optical energy by ~40 times over the output of a previously used photorefractive Bi12SiO20 crystal. Furthermore, using both a photorefractive polymer and a Bi12SiO20 crystal as the phase-conjugate mirrors, we show direct visualization and dynamic control of TRUE focus, and demonstrate fluorescence imaging in a thick turbid medium. The last part of this dissertation describes improvements in the scanning speed of a TRUE focus, using digital phase-conjugate mirrors in both transmission and reflection modes. By employing a multiplex recording of ultrasonically encoded wavefronts in transmission mode, we have accelerated the generation of multiple TRUE foci, using frequency sweeping of both ultrasound and light. With this technique, we obtained a 2-D image of a fluorescent target centered inside a turbid sample having a thickness of 2.4 lt'. Also, by gradually moving the focal position in reflection mode, we show that the TRUE focal intensity is improved, and can be continuously scanned to image fluorescent targets in a shorter time.
Ultrasound-mediated imaging in scattering media has deep roots in condensed matter physics. This book explores these roots and provides a comprehensive review of imaging with light and ultrasound, focusing mainly on recovery of mechanical contrasts. It is suitable reference material for graduate students and practitioners in the field.
Ultrasound modulated optical tomography is a developing hybrid imaging modality that combines high optical contrast and good ultrasonic resolution to image 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 mechanical properties of biological tissues, with improved detection efficiency of ultrasound-tagged photons. We also applied 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 was improved by using a Tm3+: YAG crystal of higher doping concentration (2.0-atomic%) and a double-pass pumping configuration. With the improved SHB-UOT system, we imaged absorbing, scattering, and phase contrast objects that were 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 imaged two absorbing objects embedded in the middle plane of a 32-mm thick chicken breast sample. The results suggest that the improved SHB-UOT system is one step closer to a practical optical imaging application in biological and clinical studies. Light focusing plays a central role in biomedical imaging, manipulation, and therapy. In optical scattering media such as biological tissue, light propagation is randomized by multiple scattering. Beyond one transport mean free path, where photon propagation is in the diffusive regime, direct light focusing becomes infeasible. Although various methods have been developed to overcome this optical diffusion limit, all are limited by the lack of a practical internal "guide star." Here we proposed and experimentally validated a novel concept, called Time-Reversed Ultrasonically Encoded (TRUE) optical focusing, to deliver light dynamically into any predefined location inside a scattering medium. First, diffused coherent light is encoded by an ultrasonic wave focused to a predefined location; then, the encoded component of the diffused light is time-reversed and consequently converges back to the ultrasonic focus. The ultrasonic encoding noninvasively provides a virtual internal "guide star" for the time reversal. The TRUE optical focus--dynamically defined by the ultrasonic focus--is unaffected by multiple scattering of light, which is especially desirable in biological tissue where ultrasonic scattering is ~1000 times weaker than optical scattering. Various fields, such as biomedical, colloidal, atmospheric, and ocean optics, can benefit from TRUE optical focusing. Further, the concept can be generalized for non-optical waves.
Ultrasound-mediated imaging in scattering media has deep roots in condensed matter physics. This book explores these roots and provides a comprehensive review of imaging with light and ultrasound, focusing mainly on recovery of mechanical contrasts. It is suitable reference material for graduate students and practitioners in the field.
In scattering media such as biological tissue, the heterogeneous refractive index distribution causes light to scatter, which makes the media look opaque and prevents us from focusing light beyond 1̃ mm deep inside the media to achieve optical imaging and manipulation. Hence, the ability to focus light deep inside scattering media is highly desired, and it could revolutionize biophotonics by enabling deep-tissue non-invasive high-resolution optical microscopy, optical tweezing, optogenetics, micro-surgery, and phototherapy. To break the optical diffusion limit and focus light deep inside scattering media, optical phase conjugation based wavefront shaping techniques, such as time-reversed ultrasonically encoded (TRUE) optical focusing, are being actively developed. In this dissertation, I will describe our efforts to improve the performance (speed, focusing quality and focusing depth) of optical phase conjugation for future in vivo applications. Remarkably, we have focused light through tissue-mimicking phantoms up to 96 mm thick, and through ex vivo chicken breast tissue up to 25 mm thick.
Optical imaging of soft biological tissue is highly desirable since it is nonionizing and provides sensitive contrast information which enables the detection of physiological functions and abnormalities, including potentially early cancer detection. However, due to the diffusive nature of light in soft biological tissue, it is difficult to achieve simultaneously good spatial resolution and good imaging depth with pure optical imaging modalities. This work focuses on the ultrasound-modulated optical tomography (UOT): a hybrid technique which combines the advantages of ultrasonic resolution and optical contrast. In this technique, focused ultrasound and optical radiation of high temporal coherence are simultaneously applied to soft biological tissue. The intensity of the sideband, or ultrasound "tagged" photons depends on the optical absorption in the region of interest where the ultrasound is focused. Demodulation of the optical speckle pattern yields the intensity of tagged photons for each location of the ultrasonic focal spot. Thus UOT yields an image with spatial resolution of the focused ultrasound -- typically submillimeter -- whose contrast is related to local optical absorption and the diffusive properties of light in the organ. Thus it extends all the advantages of optical imaging deep into highly scattering tissue. However lack of efficient tagged light detection techniques has so far prevented ultrasound-modulated optical tomography from achieving maturity. The signal-to-noise ratio (SNR) and imaging speed are two of the most important figures of merit and need further improvement for UOT to become widely applicable. In the first part of this work, nonlinear optics detection methods have been implemented to demodulate the "tagged" photons. The most common of these is photorefractive (PR) two wave mixing (TWM) interferometry, which is a time-domain filtering technique. When used for UOT, it is found that this approach extracts not only optical properties but also mechanical properties for the area of interest. To improve on TWM, PR four wave mixing (FWM) experiments were performed to read out only the modulated light and at the same time strongly suppressing the "untagged" light. Spectral-hole burning (SHB) in a rare-earth-ion-doped crystal has been developed for UOT more recently. Experiments in Tm3 :Y3Al5O12 (Tm:YAG) show the outstanding features of SHB: large angle acceptance (etendue), light speckle processing in parallel (insensitive to the diffusive light nature) and real-time signal collection (immune to light speckle decorrelation). With the help of advanced laser stabilization techniques, two orders of magnitude improvement of SNR have been achieved in a persistent SHB material (Pr3 :Y2SiO5) compared to Tm:YAG. Also slow light with PSHB further reduces noise in Pr:YSO UOT that is caused by polarization leakage by performing time-domain filtering.
This entry-level textbook, covering the area of tissue optics, is based on the lecture notes for a graduate course (Bio-optical Imaging) that has been taught six times by the authors at Texas A&M University. After the fundamentals of photon transport in biological tissues are established, various optical imaging techniques for biological tissues are covered. The imaging modalities include ballistic imaging, quasi-ballistic imaging (optical coherence tomography), diffusion imaging, and ultrasound-aided hybrid imaging. The basic physics and engineering of each imaging technique are emphasized. A solutions manual is available for instructors; to obtain a copy please email the editorial department at [email protected].
One of the prime limiting factors of optical imaging in biological applications is the diffusion of light by tissue, which prevents focusing at depths greater than the optical diffusion limit of ~1 mm in soft tissue. This greatly restricts the utility of optical diagnostic and therapeutic techniques, such as optogenetics, microsurgery, optical tweezing, and phototherapy of deep tissue, which require focused light in order to function. Wavefront shaping extends the depth at which optical focusing may be achieved by compensating for phase distortions induced by scattering, allowing for focusing through constructive interference. However, due to physiological motion, scattering of light in tissue is deterministic only within a brief speckle correlation time. In in vivo soft tissue, this speckle correlation is on the order of milliseconds. Because wavefront shaping relies on deterministic scattering in order to compensate for the resulting phase distortion, the wavefront must be optimized within this brief period. This presents a challenge as the speed of digital wavefront shaping has typically been limited by the relatively long time required to measure and display the optimal phase pattern due to the low speed of cameras, data transfer and processing, and spatial light modulators. In order to overcome these restrictions, wavefront shaping techniques which minimize the time required in measurement and display are therefore vital. In this dissertation, I will describe our efforts to improve the speed of wavefront shaping without sacrificing the performance of the systems. To this end, we have successfully developed several systems which are capable of full-phase wavefront shaping with latencies of 9 ms or less. In addition, we report an all-digital alignment compensation protocol, which may be used to obtain optimal alignment in digital optical phase conjugation systems, a key component when acquiring the best possible focusing performance.
The use of light for probing and imaging biomedical media is promising for the development of safe, noninvasive, and inexpensive clinical imaging modalities with diagnostic ability. The advent of ultrafast lasers has enabled applications of nonlinear optical processes, which allow deeper imaging in biological tissues with higher spatial resolution. This book provides an overview of emerging novel optical imaging techniques, Gaussian beam optics, light scattering, nonlinear optics, and nonlinear optical tomography of tissues and cells. It consists of pioneering works that employ different linear and nonlinear optical imaging techniques for deep tissue imaging, including the new applications of single- and multiphoton excitation fluorescence, Raman scattering, resonance Raman spectroscopy, second harmonic generation, stimulated Raman scattering gain and loss, coherent anti-Stokes Raman spectroscopy, and near-infrared and mid-infrared supercontinuum spectroscopy. The book is a comprehensive reference of emerging deep tissue imaging techniques for researchers and students working in various disciplines.
"In optical imaging, the depth and resolution are limited due to scattering. Unlike light, scattering of an ultrasound waves in tissue is negligible. Hybrid imaging methods such as ultrasound modulated optical tomography (UOT) use the advantages of both modalities. UOT tags light by inducing phase change caused by modulating the local index of refraction of the medium. The challenge in UOT is detecting the weak signal. The displacement induced by the acoustic radiation force (ARF) is another ultrasound effect that can be utilized to tag the light. It induces greater phase change, resulting in a stronger signal. Moreover, the absorbed acoustic energy generates heat, resulting in a change in the index of refraction and a strong phase change. Since the speckle pattern is governed by the phase of the interfering scattered waves speckle pattern analysis can obtain information about local displacement and temperature changes in the tissue. We have presented a model to simulate the insonation processes. Simulation results based on fixed-particle Monte Carlo and experimental results show that the signal acquired by utilizing ARF is stronger compared to UOT. The introduced mean irradiance change (MIC) signal reveals both thermal and mechanical effects of the focused ultrasound wave in different timescales. Simulation results suggest that variation in the MIC signal can be used to generate a displacement image of the medium. In addition to the displacement correlated image, the MIC signal can provide images based on the morphology of the tissue. The MIC signal can provide for tumor detection in a healthy tissue"--Author's abstract.