Puxiang Lai

Optical focusing deep inside dynamic scattering media with near-infrared time-reversed ultrasonically encoded (TRUE) light.

Focusing light deep inside living tissue has not been achieved despite its promise to play a central role in biomedical imaging, optical manipulation and therapy. To address this challenge, internal-guide-star-based wavefront engineering techniques--for example, time-reversed ultrasonically encoded (TRUE) optical focusing--were developed. The speeds of these techniques, however, were limited to no greater than 1 Hz, preventing them from in vivo applications. Here we improve the speed of optical focusing deep inside scattering media by two orders of magnitude, and focus diffuse light inside a dynamic scattering medium having a speckle correlation time as short as 5.6 ms, typical of living tissue. By imaging a target, we demonstrate the first focusing of diffuse light inside a dynamic scattering medium containing living tissue. Since the achieved focusing speed approaches the tissue decorrelation rate, this work is an important step towards in vivo deep tissue noninvasive optical imaging, optogenetics and photodynamic therapy.

Ultrasonically encoded wavefront shaping for focusing into random media

Focusing light into opaque random or scattering media such as biological tissue is a much sought-after goal for biomedical applications such as photodynamic therapy, optical manipulation, and photostimulation. However, focusing with conventional lenses is restricted to one transport mean free path in scattering media, limiting both optical penetration depth and resolution. Focusing deeper is possible by using optical phase conjugation or wavefront shaping to compensate for the scattering. For practical applications, wavefront shaping offers the advantage of a robust optical system that is less sensitive to optical misalignment. Here, the phase of the incident light is spatially tailored using a phase-shifting array to pre-compensate for scattering. The challenge, then, is to determine the phase pattern which allows light to be optimally delivered to the target region. Optimization algorithms are typically employed for this purpose, with visible particles used as targets to generate feedback. However, using these particles is invasive, and light delivery is limited to fixed points. Here, we demonstrate a method for non-invasive and dynamic focusing, by using ultrasound encoding as a virtual guide star for feedback to an optimization algorithm. The light intensity at the acoustic focus was increased by an order of magnitude. This technique has broad biomedical applications, such as in optogenetics or photoactivation of drugs.Phase distortions due to scattering in random media restrict optical focusing beyond one transport mean free path. However, scattering can be compensated for by applying a correction to the illumination wavefront using spatial light modulators. One method of obtaining the wavefront correction is by iterative determination using an optimization algorithm. In the past, obtaining a feedback signal required either direct optical access to the target region, or invasive embedding of molecular probes within the random media. Here, we propose using ultrasonically encoded light as feedback to guide the optimization dynamically and non-invasively. In our proof-of-principle demonstration, diffuse light was refocused to the ultrasound focal zone, with a focus-to-background ratio of more than one order of magnitude after 600 iterations. With further improvements, especially in optimization speed, the proposed method should find broad applications in deep tissue optical imaging and therapy.

Focused fluorescence excitation with time-reversed ultrasonically encoded light and imaging in thick scattering media

Scattering dominates light propagation in biological tissue, and therefore restricts both resolution and penetration depth in optical imaging within thick tissue. As photons travel into the diffusive regime-typically 1 mm beneath human skin, their trajectories transition from ballistic to diffusive due to increased number of scattering events, which makes it impossible to focus, much less track, photon paths. Consequently, imaging methods that rely on controlled light illumination are ineffective in deep tissue. This problem has recently been addressed by a novel method capable of dynamically focusing light in thick scattering media via time reversal of ultrasonically encoded (TRUE) diffused light. Here, using photorefractive materials as phase conjugate mirrors, we show a direct visualization and dynamic control of optical focusing with this light delivery method, and demonstrate its application for focused fluorescence excitation and imaging in thick turbid media. These abilities are increasingly critical to understanding the dynamic interactions of light with biological matter and processes at different system levels, as well as their applications for biomedical diagnosis and therapy.

Reflection-mode time-reversed ultrasonically encoded optical focusing into turbid media

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 in biological tissue

We report an experimental investigation of time-reversed ultrasonically encoded optical focusing in biological tissue. This technology combines the concepts of optical phase conjugation and ultrasound modulation of diffused coherent light. The ultrasonically encoded (or tagged) diffused light from a tissue sample is collected in reflection mode and interferes with a reference light in a photorefractive crystal (used as a phase conjugation mirror) to form a hologram. Then a time-reversed copy of the tagged light is generated and traces back the original trajectories to the ultrasonic focus inside the tissue sample. With our current setup, we can achieve a maximum penetration depth of 5 mm in a chicken breast sample and image optical contrasts within a tissue sample with a spatial resolution approximately equaling 1/√2 of the ultrasound focal diameter.

REAL-TIME MONITORING OF HIGH-INTENSITY FOCUSED ULTRASOUND LESION FORMATION USING ACOUSTO-OPTIC SENSING

High-intensity focused ultrasound (HIFU) is a promising modality that is used to noninvasively ablate soft tissue tumors. Nevertheless, real-time treatment monitoring with diagnostic ultrasound still poses a significant challenge since tissue necrosis, in the absence of cavitation or boiling, provides little acoustic contrast with normal tissue. In comparison, the optical properties of tissue are significantly altered accompanying lesion formation. A photorefractive crystal-based acousto-optic (AO) sensing system that uses a single HIFU transducer to simultaneously generate tissue necrosis and pump the AO interaction is used to monitor the real-time optical changes associated with thermal lesions induced in chicken breast ex vivo. It is found that the normalized change in AO response increases proportionally with the volume of necrosis. This study demonstrates AO sensing can identify the onset and growth of lesion formation in real time and, when used as feedback to guide exposures, results in more predictable lesion formation.

Ultrasound-modulated optical tomography at new depth

Ultrasound-modulated optical tomography (UOT) has the potential to reveal optical contrast deep inside soft biological tissues at an ultrasonically determined spatial resolution. The optical imaging depth reported so far has, however, been limited, which prevents this technique from broader applications. Our latest experimental exploration has pushed UOT to an unprecedented imaging depth. We developed and optimized a UOT system employing a photorefractive crystal-based interferometer. A large aperture optical fiber bundle was used to enhance the efficiencies for diffuse light collection and photorefractive two-wave-mixing. Within the safety limits for both laser illumination and ultrasound modulation, the system has attained the ability to image through a tissue-mimicking phantom of 9.4 cm in thickness, which has never been reached previously by UOT.

Time-reversed ultrasonically encoded optical focusing into tissue-mimicking media with thickness up to 70 mean free paths

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.

Dependence of optical scattering from Intralipid in gelatin-gel based tissue-mimicking phantoms on mixing temperature and time

Abstract. Intralipid is widely used as an optical scattering agent in tissue-mimicking phantoms. Accurate control when using Intralipid is critical to match the optical diffusivity of phantoms to the prescribed value. Currently, most protocols of Intralipid-based hydrogel phantom fabrication focus on factors such as Intralipid brand and concentration. In this note, for the first time to our knowledge, we explore the dependence of the optical reduced scattering coefficient (at 532 nm optical wavelength) on the temperature and the time of mixing Intralipid with gelatin-water solution. The studied samples contained 1% Intralipid and were measured with oblique-incidence reflectometry. It was found that the reduced scattering coefficient increased when the Intralipid-gelatin-water mixture began to solidify at room temperature. For phantoms that had already solidified completely, the diffusivity was shown to be significantly influenced by the temperature and the duration of the mixing course. The dependence of the measured diffusivity on the mixing conditions was confirmed by experimental observations. Moreover, the mechanism behind the dependence behavior is discussed.

Energy enhancement in time-reversed ultrasonically encoded optical focusing using a photorefractive polymer.

Time-reversed ultrasonically encoded (TRUE) optical focusing achieves light focusing into scattering media beyond one transport mean free path, which is desirable in biomedical optics. However, the focused optical energy needs to be increased for broad applications. Here, we report the use of a photorefractive polymer (PRP) as the phase conjugate mirror in TRUE optical focusing. The PRP boosted the focused optical energy by ~40 times in comparison to the previously used photorefractive Bi12SiO20 crystal. As a result, we successfully imaged absorbing objects embedded in the middle plane of a tissue-mimicking phantom having an optical thickness of 120 scattering mean free paths.