Optical-resolution photoacoustic endomicroscopy in vivo.
ABSTRACT: Optical-resolution photoacoustic microscopy (OR-PAM) has become a major experimental tool of photoacoustic tomography, with unique imaging capabilities for various biological applications. However, conventional imaging systems are all table-top embodiments, which preclude their use in internal organs. In this study, by applying the OR-PAM concept to our recently developed endoscopic technique, called photoacoustic endoscopy (PAE), we created an optical-resolution photoacoustic endomicroscopy (OR-PAEM) system, which enables internal organ imaging with a much finer resolution than conventional acoustic-resolution PAE systems. OR-PAEM has potential preclinical and clinical applications using either endogenous or exogenous contrast agents.
Project description:Multimode fibres (MMFs) are becoming increasingly attractive in optical endoscopy as they promise to enable unparallelled miniaturisation, spatial resolution and cost. However, high-speed imaging with wavefront shaping has been challenging. Here, we report the development of a video-rate dual-modal photoacoustic (PA) and fluorescence microscopy probe with a high-speed digital micromirror device (DMD) towards forward-viewing endomicroscopy. Optimal DMD patterns were obtained using a real-valued intensity transmission matrix algorithm to raster-scan a 1.5 μ m-diameter focused beam at the distal fibre tip for imaging. The PA imaging speed and spatial resolution were varied from ∼ 2 to 57 frames per second and from 1.7 to 3 μ m, respectively. Further, high-fidelity PA images of carbon fibres and mouse red blood cells were acquired at unprecedented speed. The capability of dual-modal imaging was demonstrated with phantoms. We anticipate that with further miniaturisation of the ultrasound detector, this probe could be integrated into medical needles to guide minimally invasive procedures.
Project description:Photoacoustic microscopy (PAM) is uniquely positioned for biomedical applications because of its ability to visualize optical absorption contrast in vivo in three dimensions. Here we propose motionless volumetric spatially invariant resolution photoacoustic microscopy (SIR-PAM). To realize motionless volumetric imaging, SIR-PAM combines two-dimensional Fourier-spectrum optical excitation with single-element depth-resolved photoacoustic detection. To achieve spatially invariant lateral resolution, propagation-invariant sinusoidal fringes are generated by a digital micromirror device. Further, SIR-PAM achieves 1.5 times finer lateral resolution than conventional PAM. The superior performance was demonstrated in imaging both inanimate objects and animals in vivo with a resolution-invariant axial range of 1.8?mm, 33 times the depth of field of the conventional PAM counterpart. Our work opens new perspectives for PAM in biomedical sciences.Photoacoustic microscopy allows for label-free 3D in vivo imaging by detecting the acoustic response of a photoexcited material. Here, Yang et. al use a digital-micromirror-device based structured illumination scheme to both improve resolution and greatly increase the depth of field, enabling 3D volumetric imaging.
Project description:BACKGROUND:Focal therapy for prostate cancer has been proposed as an alternative treatment to whole-gland therapies in selected men to diminish side effects in localized prostate cancer. As nowadays imaging cannot offer complete prostate cancer disease characterization, multicore systematic biopsies are recommended (transrectal or transperineal). Optical imaging techniques such as confocal laser endomicroscopy and optical coherence tomography allow in vivo, high-resolution imaging. Moreover, they can provide real-time visualization and analysis of tissue and have the potential to offer additive diagnostic information. OBJECTIVE:This study has 2 separate primary objectives. The first is to assess the technical feasibility and safety of in vivo focal imaging with confocal laser endomicroscopy and optical coherence tomography. The second is to identify and define characteristics of prostate cancer and normal prostate tissue in confocal laser endomicroscopy and optical coherence tomography imaging by comparing these images with the corresponding histopathology. METHODS:In this prospective, in vivo feasibility study, needle-based confocal laser endomicroscopy and optical coherence tomography imaging will be performed before transperineal template mapping biopsy or radical prostatectomy. First, confocal laser endomicroscopy and optical coherence tomography will be performed in 4 patients (2 for each imaging modality) undergoing transperineal template mapping biopsy to assess the feasibility and safety of confocal laser endomicroscopy and optical coherence tomography. If proven to be safe and feasible, confocal laser endomicroscopy and optical coherence tomography will be performed in 10 patients (5 for each imaging modality) undergoing radical prostatectomy. Confocal laser endomicroscopy and optical coherence tomography images will be analyzed by independent, blinded observers. Confocal laser endomicroscopy- and optical coherence tomography-based qualitative and quantitative characteristics and histopathology will be compared. The study complies with the IDEAL (Idea, Development, Exploration, Assessment, Long-term study) stage 2a recommendations. RESULTS:At present, the study is enrolling patients and results and outcomes are expected in 2019. CONCLUSIONS:Confocal laser endomicroscopy and optical coherence tomography are promising optical imaging techniques that can visualize and analyze tissue structure, possible tumor grade, and architecture in real time. They can potentially provide real-time, high-resolution microscopic imaging and tissue characteristics of prostate cancer in conjunction with magnetic resonance imaging or transrectal ultrasound fusion-guided biopsy procedures. This study will provide insight into the feasibility and tissue-specific characteristics of confocal laser endomicroscopy and optical coherence tomography for real-time optical analysis of prostate cancer. TRIAL REGISTRATION:ClinicalTrials.gov NCT03253458; https://clinicaltrials.gov/ct2/show/NCT03253458 (Archived by WebCite at http://www.webcitation.org/6z9owM66B). REGISTERED REPORT IDENTIFIER:RR1-10.2196/9813.
Project description:Photoacoustic tomography (PAT) is a molecular imaging technology. Unlike conventional reporter gene imaging, which is usually based on fluorescence, photoacoustic reporter gene imaging relies only on optical absorption. This work demonstrates several key merits of PAT using lacZ, one of the most widely used reporter genes in biology. We show that the expression of lacZ can be imaged by PAT as deep as 5.0 cm in biological tissue, with resolutions of ?1.0 mm and ?0.4 mm in the lateral and axial directions, respectively. We further demonstrate non-invasive, simultaneous imaging of a lacZ-expressing tumor and its surrounding microvasculature in vivo by dual-wavelength acoustic-resolution photoacoustic microscopy (AR-PAM), with a lateral resolution of 45 µm and an axial resolution of 15 µm. Finally, using optical-resolution photoacoustic microscopy (OR-PAM), we show intra-cellular localization of lacZ expression, with a lateral resolution of a fraction of a micron. These results suggest that PAT is a complementary tool to conventional optical fluorescence imaging of reporter genes for linking biological studies from the microscopic to the macroscopic scales.
Project description:The temperature-dependent property of the Grueneisen parameter has been employed in photoacoustic imaging mainly to measure tissue temperature. Here we explore this property using a different approach and develop Grueneisen relaxation photoacoustic microscopy (GR-PAM), a technique that images nonradiative absorption with confocal optical resolution. GR-PAM sequentially delivers two identical laser pulses with a microsecond-scale time delay. The first laser pulse generates a photoacoustic signal and thermally tags the in-focus absorbers. When the second laser pulse excites the tagged absorbers within the thermal relaxation time, a photoacoustic signal stronger than the first one is produced, owing to the temperature dependence of the Grueneisen parameter. GR-PAM detects the amplitude difference between the two colocated photoacoustic signals, confocally imaging the nonradiative absorption. We greatly improved axial resolution from 45???m to 2.3???m and, at the same time, slightly improved lateral resolution from 0.63???m to 0.41???m. In addition, the optical sectioning capability facilitates the measurement of the absolute absorption coefficient without fluence calibration.
Project description:High-speed optical-resolution photoacoustic microscopy (OR-PAM), integrating the merits of high spatial resolution and fast imaging acquisition, can observe dynamic processes of the optical absorption-based molecular specificities. However, it remains challenging for the evaluation to morphological and physiological parameters that are closely associated with photoacoustic spectrum due to the inadequate ultrasonic frequency response of the routinely-employed piezoelectric transducer. By utilizing the galvanometer for fast optical scanning and our previously-developed surface plasmon resonance sensor as an unfocused broadband ultrasonic detector, high-speed spectroscopic photoacoustic imaging was accessed in the OR-PAM system, achieving an acoustic bandwidth of ∼125 MHz and B-scan rate at ∼200 Hz over a scanning range of ∼0.5 mm. Our system demonstrated the dynamic imaging of the moving phantoms' structures and the simultaneous characterization of their photoacoustic spectra over time. Further, fast volumetric imaging and spectroscopic analysis of microanatomic features of a zebrafish eye ex vivo was obtained label-freely.
Project description:Compared with single-focus optical-resolution photoacoustic microscopy (OR-PAM), multifocal OR-PAM utilizes both multifocal optical illumination and an ultrasonic array transducer, significantly increasing the imaging speed. A reflection-mode multifocal OR-PAM system based on a microlens array that provides multiple foci as well as an ultrasonic array transducer that receives the excited photoacoustic waves from all foci simultaneously is presented. Using a customized microprism to reflect the incident laser beam to the microlens array, the multiple optical foci are aligned confocally with the focal zone of the ultrasonic array transducer. Experiments show the reflection-mode multifocal OR-PAM is capable of imaging microvessels in vivo, and it can image a 6×5×2.5??mm³ volume at 16 ?m lateral resolution in ?2.5??min, which was limited by the signal multiplexing ratio and laser pulse repetition rate.
Project description:Photoacoustic microscopy (PAM) is an attractive imaging tool complementary to established optical microscopic modalities by providing additional molecular specificities through imaging optical absorption contrast. While the development of optical resolution photoacoustic microscopy (ORPAM) offers high lateral resolution, the acoustically-determined axial resolution is limited due to the constraint in ultrasonic detection bandwidth. ORPAM with isometric spatial resolution along both axial and lateral direction is yet to be developed. Although recently developed sophisticated optical illumination and reconstruction methods offer improved axial resolution in ORPAM, the image acquisition procedures are rather complicated, limiting their capabilities for high-speed imaging and being easily integrated with established optical microscopic modalities. Here we report an isometric ORPAM based on an optically transparent micro-ring resonator ultrasonic detector and a commercial inverted microscope platform. Owing to the superior spatial resolution and the ease of integrating our ORPAM with established microscopic modalities, single cell imaging with extrinsic fluorescence staining, intrinsic autofluorescence, and optical absorption can be achieved simultaneously. This technique holds promise to greatly improve the accessibility of PAM to the broader biomedical researchers.
Project description:An innovative imaging tool enables in vivo high-resolution visualization of deep brain structures and functions over large volume. Optical deep-brain imaging in vivo at high resolution has remained a great challenge over the decades. Two-photon endomicroscopy provides a minimally invasive approach to image buried brain structures, once it is integrated with a gradient refractive index (GRIN) lens embedded in the brain. However, its imaging resolution and field of view are compromised by the intrinsic aberrations of the GRIN lens. Here, we develop a two-photon endomicroscopy by adding adaptive optics based on direct wavefront sensing, which enables recovery of diffraction-limited resolution in deep-brain imaging. A new precompensation strategy plays a critical role to correct aberrations over large volumes and achieve rapid random-access multiplane imaging. We investigate the neuronal plasticity in the hippocampus, a critical deep brain structure, and reveal the relationship between the somatic and dendritic activity of pyramidal neurons.
Project description:Photoacoustic imaging (PAI) is a noninvasive imaging tool to visualize optical absorbing contrast agents. Due to high ultrasonic resolution and superior optical sensitivity, PAI can be used to monitor nanoparticle-mediated cancer therapy. The current study synthesized Food and Drug Administration-approved Prussian blue (PB) in the form of nanoparticles (NPs) with the peak absorption at 712?nm for photoacoustically imaging tumor-bearing mouse models. To monitor PB NPs from the background tissue in vivo, we also developed a new 700-nm-region stimulated Raman scattering (SRS) source (pulse energy up to 200?nJ and repetition rate up to 50?kHz) and implemented optical-resolution photoacoustic microscopy (OR-PAM). The SRS-assisted OR-PAM system was able to monitor PB NPs in the tumor model with micrometer resolution. Due to strong light absorption at 712?nm, the developed SRS light yielded a two-fold higher contrast from PB NPs, in comparison with a 532-nm pumping source. The proposed laser source involved cost-effective and simple system implementation along with high compatibility with the fiber-based OR-PAM system. The study highlights the OR-PAM system in conjunction with the tunable-color SRS light source as a feasible tool to assist NP-mediated cancer therapy.