Optoacoustic imaging, or photoacoustic imaging, is insensitive to photon scattering within biological tissue and, unlike conventional optical imaging methods, makes high-resolution optical visualization deep within tissue possible. Recent advances in laser technology, detection strategies and inversion techniques have led to significant improvements in the capabilities of optoacoustic systems. A key empowering feature is the development of video-rate multispectral imaging in two and three dimensions, which offers fast, spectral differentiation of distinct photoabsorbing moieties. We review recent advances and capabilities in the technology and its corresponding emerging biological and clinical applications.

Advances in real-time multispectral optoacoustic imaging and its applications

Photoacoustic microscopy (PAM) is a hybrid in vivo imaging technique that acoustically detects optical contrast via the photoacoustic effect. Unlike pure optical microscopic techniques, PAM takes advantage of the weak acoustic scattering in tissue and thus breaks through the optical diffusion limit (~1 mm in soft tissue). With its excellent scalability, PAM can provide high-resolution images at desired maximum imaging depths up to a few millimeters. Compared with backscattering-based confocal microscopy and optical coherence tomography, PAM provides absorption contrast instead of scattering contrast. Furthermore, PAM can image more molecules, endogenous or exogenous, at their absorbing wavelengths than fluorescence-based methods, such as wide-field, confocal, and multi-photon microscopy. Most importantly, PAM can simultaneously image anatomical, functional, molecular, flow dynamic and metabolic contrasts in vivo. Focusing on state-of-the-art developments in PAM, this Review discusses the key features of PAM implementations and their applications in biomedical studies.

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Photoacoustic microscopy: Photoacoustic microscopy

In the last 25 years, optical coherence tomography (OCT) has advanced to be one of the most innovative and most successful translational optical imaging techniques, achieving substantial economic impact as well as clinical acceptance. This is largely owing to the resolution improvements by a factor of 10 to the submicron regime and to the imaging speed increase by more than half a million times to more than 5 million A-scans per second, with the latter one accomplished by the state-of-the-art swept source laser technologies that are reviewed in this article. In addition, parallelization of OCT detection, such as line-field and full-field OCT, has shortened the acquisition time even further by establishing quasi-akinetic scanning. Besides the technical improvements, several functional and contrast-enhancing OCT applications have been investigated, among which the label-free angiography shows great potential for future studies. Finally, various multimodal imaging modalities with OCT incorporated are reviewed, in that these multimodal implementations can synergistically compensate for the fundamental limitations of OCT when it is used alone.

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Optical coherence tomography today: speed, contrast, and multimodality.

Photoacoustic tomography (PAT) is an emerging imaging modality that shows great potential for preclinical research and clinical practice. As a hybrid technique, PAT is based on the acoustic detection of optical absorption from either endogenous chromophores, such as oxy-hemoglobin and deoxy-hemoglobin, or exogenous contrast agents, such as organic dyes and nanoparticles. Because ultrasound scatters much less than light in tissue, PAT generates high-resolution images in both the optical ballistic and diffusive regimes. Over the past decade, the photoacoustic technique has been evolving rapidly, leading to a variety of exciting discoveries and applications. This review covers the basic principles of PAT and its different implementations. Strengths of PAT are highlighted, along with the most recent imaging results.

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Photoacoustic tomography: principles and advances.

Photoacoustic tomography (PAT) has become one of the fastest growing fields in biomedical optics. Unlike pure optical imaging, such as confocal microscopy and two-photon microscopy, PAT employs acoustic detection to image optical absorption contrast with high-resolution deep into scattering tissue. So far, PAT has been widely used for multiscale anatomical, functional, and molecular imaging of biological tissues. We focus on PAT’s basic principles, major implementations, imaging contrasts, and recent applications.

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Tutorial on photoacoustic tomography

Elasto-optical refractive index modulation due to photoacoustic initial pressure transients produced significant reflection of a probe beam when the absorbing interface had an appreciable refractive index difference This effect was harnessed in a new form of non-contact optical resolution photoacoustic microscopy called photoacoustic remote sensing microscopy A non-interferometric system architecture with a low-coherence probe beam precludes detection of surface oscillations and other phase-modulation phenomenon The probe beam was confocal with a scanned excitation beam to ensure detection of initial pressure-induced intensity reflections at the subsurface origin where pressures are largest Phantom studies confirmed signal dependence on optical absorption, index contrast and excitation fluence In vivo imaging of superficial microvasculature and melanoma tumors was demonstrated with ~27±05 μm lateral resolution A new design for a photoacoustic microscope capable of high-quality, real-time in vivo imaging has been developed by scientists in Canada Parsin Hajireza and co-workers from the University of Alberta and the company Illumisonics report that, unlike other designs, their approach does not rely on interferometric detection of photoacoustic stress, which can be problematic Instead, it involves making time-varying intensity measurements of the reflection of a probe beam from the sample A high signal-to-noise ratio and a working distance of 25 centimetres between the sample and the system's objective lens are achievable The researchers demonstrate the potential of their scheme for biomedical applications by using to perform in vivo imaging of microvasculature and melanoma tumours in chicken embryos with a spatial resolution of 27 micrometres

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Non-interferometric photoacoustic remote sensing microscopy

In this Letter, the capability of label-free fiber-based optical-resolution photoacoustic microscopy is demonstrated. This real-time imaging system takes advantage of image-guide fibers and a unique fiber laser. The 800 μm image-guide consists of 30,000 individual single-mode fibers in a bundle and the diode-pumped, pulsed Ytterbium fiber laser is utilized to perform up to 600 kHz repetition rate. Phantom studies indicate 7 μm resolution. The proposed setup keeps many of the powerful properties of previous tabletop OR-PAM systems, but also offers a submillimeter probe footprint and high flexibility due to the nature of the image-guide. This system could have significant clinical impact for endoscopic applications where the thin fiber can be inserted into the body.

Label-free in vivo fiber-based optical-resolution photoacoustic microscopy

In this paper a multi-wavelength optical-resolution photoacoustic microscopy (OR-PAM) system using stimulated Raman scattering is demonstrated for both phantom and in vivo imaging. A 1-ns pulse width ytterbium-doped fiber laser is coupled into a single-mode polarization maintaining fiber. Discrete Raman-shifted wavelength peaks extending to nearly 800 nm are generated with pulse energies sufficient for OR-PAM imaging. Bandpass filters are used to select imaging wavelengths. A dual-mirror galvanometer system was used to scan the focused outputs across samples of carbon fiber networks, 200μm dye-filled tubes, and Swiss Webster mouse ears. Photoacoustic signals were collected in transmission mode and used to create maximum amplitude projection C-scan images. Double dye experiments and in vivo oxygen saturation estimation confirmed functional imaging potential.

In-Vivo functional optical-resolution photoacoustic microscopy with stimulated Raman scattering fiber-laser source.

In this paper a new class of optical Fabry-Perot-based ultrasound detectors using low acoustic impedance glancing angle deposited (GLAD) films is demonstrated. GLAD is a single-step physical vapor-deposition (PVD) technique used to fabricate porous nanostructured thin films. Using titanium dioxide (TiO(2)), a transparent semiconductor with a high refractive index (n = 2.4), the GLAD technique can be employed to fabricate samples with tailored nano-porosity, refractive index periodicities, and high Q-factor reflectance spectra. The average acoustic impedance of the porous films is lower than bulk materials which will improve acoustic coupling, especially for high acoustic frequencies. For this work, two filters with high reflection in the C-band range and high transparency in the visible range (~80%) using GLAD films were fabricated. A 23 µm Parylene C layer was sandwiched between these two GLAD films in order to form a GLAD Fabry Perot Interferometer (GLAD-FPI). A high speed tunable continuous wavelength C-band laser was focused at the FPI and the reflection was measured using a high speed photodiode. The ultrasound pressure modulated the optical thickness of the FPI and hence its reflectivity. The fabricated sensor was tested using a 10 MHz unfocused transducer. The ultrasound transducer was calibrated using a hydrophone. The minimum detectable acoustic pressure was measured as 80 ± 20 Pa and the -3dB bandwidth was measured to be 18 MHz. This ultra-sensitive sensor can be an alternative to piezoelectric ultrasound transducers for any techniques in which ultrasound waves need to be detected including ultrasonic and photoacoustic imaging modalities. We demonstrate our GLAD-FPI for photoacoustic signal detection in optical-resolution photoacoustic microscopy (OR-PAM). To the best of our knowledge, this is the first time that a FPI fabricated using the GLAD method has been used for ultra-sensitive ultrasound detection.

Glancing angle deposited nanostructured film Fabry-Perot etalons for optical detection of ultrasound

Optical-resolution photoacoustic microscopy (OR-PAM) is capable of achieving optical-absorption-contrast images with micron-scale spatial resolution. Previous OR-PAM systems have been frame-rate limited by mechanical scanning speeds and laser pulse repetition rate (PRR). We demonstrate OR-PAM imaging using a diode-pumped nanosecond-pulsed Ytterbium-doped 532-nm fiber laser with PRR up to 600 kHz. Combined with fast-scanning mirrors, our proposed system provides C-scan and 3D images with acquisition frame rate of 4 frames per second (fps) or higher, two orders of magnitude faster than previously published systems. High-contrast images of capillary-scale microvasculature in a live Swiss Webster mouse ear with ~6-µm optical lateral spatial resolution are demonstrated.

Parsin Hajireza

Papers

Non-interferometric photoacoustic remote sensing microscopy

Label-free in vivo fiber-based optical-resolution photoacoustic microscopy

In-Vivo functional optical-resolution photoacoustic microscopy with stimulated Raman scattering fiber-laser source.

Glancing angle deposited nanostructured film Fabry-Perot etalons for optical detection of ultrasound

In vivo near-realtime volumetric optical-resolution photoacoustic microscopy using a high-repetition-rate nanosecond fiber-laser