KEYWORDS: Sensors, Signal detection, Optoacoustics, Ultrasonography, Capacitance, Signal to noise ratio, In vivo imaging, Image processing, Objectives, Design and modelling
A wideband sensitive needle ultrasound sensor based on a polarized PVDF-TrFE copolymer piezoelectric film has been developed, which is capable of providing a noise equivalent pressure of 14 Pa and a uniform frequency response ranging from 1 to 25 MHz. Its high sensitivity (1.6 μV / Pa) and compact size were achieved by capitalizing on the large electromechanical coupling coefficient of PVDF-TrFE and minimizing parasitic capacitance in a two-stage amplifier structure. The detection sensitivity of the newly designed sensor outperformed commercially available hydrophones with an equivalent sensing element area by a factor of 9. The sensor has been successfully integrated into a light scanning optoacoustic microscopy (OAM) system with a limited working space. Submicrometer resolution images were subsequently attained from living mice without employing signal averaging. The miniature sensor design can readily be integrated into various OAM systems and further facilitate multimodal imaging system implementations.
KEYWORDS: Tumors, Tissues, Skin, Algorithm development, 3D image processing, Angiography, 3D image reconstruction, Optoacoustics, Reconstruction algorithms, In vivo imaging
We demonstrate the opportunities of the developed 3D optoacoustic image processing algorithm to characterize numerically the vasculature parameters in different applications including monitoring of tumor angiogenesis and assessing skin aging.
The possibilities of optoacoustic microscopy for comparison of vascular network of different tumor models as well as for investigation of tumor vessels response to radiation therapy were demonstrated.
KEYWORDS: Ferroelectric polymers, Spherical lenses, Antennas, Optoacoustics, 3D image processing, Sensors, In vivo imaging, Algorithm development, Signal detection, Reconstruction algorithms
We developed high-density spherical matrix array based on polyvinylidene difluoride films. Ultrawide bandwidth (0.3-38 MHz) and sub-millimeter sized elements enabled non-invasive cerebrovascular imaging of adult mouse with ~60 μm resolution.
Using optoacoustic microscopy, a radiation-induced increase in the fragmentation of experimental tumor small vessels, as well as the formation of large hemoglobin-containing structures were revealed within first days after treatment.
Our work was devoted to the experimental comparison of two ultra-wideband detectors based on PVDF piezofilms of different thickness demonstrating different quality of optoacoustic imaging of vessels in tumor and normal tissues.
We present the results of combined fluorescence and optoacoustic monitoring of tumor treatment using novel photoactivatable multi-inhibitor liposomes with BPD and Irinotecan providing a synergetic effect of PDT and chemotherapeutic impact.
We present a fast, multispectral acoustic resolution optoacoustic microscope using a new burst-mode triggering scheme. Three pulsed laser sources are combined to retrieve spectral images across large field-of-views extending over 25mm by 25mm at 28μm lateral and 14μm axial resolution with an overfly scan of a few minutes. Highly sensitive PVDF transducer allows detection of structures 3.8mm below the human skin surface with per pulse energies of only 25μJ. The newly developed system overcomes limitations of previously reported scanning optoacoustic microscopy and mesoscopy implementations, offering a major leap forward in terms of clinical usability, laser safety, effective penetration depth and spectral unmixing capabilities.
Development of a number of diseases is accompanied by changes in the blood vessels’ structure and the investigation of tissue vascular pattern remains one of the most essential problems in experimental and clinical medicine. Hybrid methods of optoacoustic (OA) imaging enable label-free optical-contrast angiography at optical penetration depths with ultrasonic resolution. We used OA to study vascular network of experimental tumor during growth and after treatment as well as to study the dynamics of blood content of human skin during functional tests. Optoacoustic angiography was performed using raster-scan system in reflection mode with 532 nm laser source and wideband PVDF detector. The sensitivity of the system allowed to visualize 50 µm blood vessels at up to 2.1 mm depth. Minimally detected diameter of blood vessel located at the surface of a soft tissue was 15 µm. Imaging of colon tumor models CT26 and HT-29 revealed peculiarities of vascular system development. Irradiation-induced increase of small tumor vessels segments number and parallel decrease in the number of large hemoglobin-containing structures were demonstrated. The fraction of blood-filled vessels of the human skin was assessed during cuff occlusion and temporarily filling with blood became clearly visible on OA images. To study the effects of local mechanical compression on human skin vasculature we varied external pressure and revealed the gradual drop in OA signal from blood vessels. We demonstrated the possibilities of raster-scan angiography for in vivo analysis of vessels structure, for monitoring of neoangiogenesis and for dynamical investigation of blood content under external actions.
Optoacoustic (OA) tomography has recently demonstrated ultrafast imaging rates, owing to its unique ability to capture volumetric image data with a single nanosecond-duration flash of light. The achievable spatial resolution of volumetric tomography is however limited by the narrow bandwidth and lack of scalability of the piezo-composite arrays currently employed for OA signal detection. Here we report on the first implementation of high-density spherical matrix array technology based on flexible polyvinylidene difluoride films featuring ultrawideband (38 MHz) sub-millimeter sized elements thus enabling volumetric imaging with ~35 μm resolution, superior image fidelity and excellent signal to noise performance permitting real-time imaging of deep tissues without employing signal averaging. We demonstrate real-time volumetric in vivo imaging capabilities of the new approach by performing cerebral angiography in mice. The new technology leverages the true potential of OA for high spatio-temporal resolution visualization of rapid biodynamics.
Cerebrovascular imaging of rodents is one of the trending applications of optoacoustics aimed at studying brain activity and pathology. Imaging of deep brain structures is often hindered by sub-optimal arrangement of the light delivery and acoustic detection systems. In our work we revisit the physics behind opto-acoustic signal generation and perform theoretical evaluation of optimal laser wavelengths with an aim of enabling whole brain cerebrovascular optoacoustic angiography in small rodents. A comprehensive model based on diffusion approximation was developed to simulate optoacoustic signal generation using optical and acoustic parameters closely mimicking a typical murine brain and surrounding tissues. The model revealed three characteristic wavelength ranges in the visible and near-infrared spectra optimally suited for imaging cerebral vasculature of different size and depth. The theoretical conclusions are confirmed by numerical simulations while in vivo imaging experiments further validated the capacity for accurately resolving brain vasculature at depths ranging between 0.7 and 7 mm.
We report on a comprehensive theoretical analysis of the optimal wavelengths for in vivo optoacoustic angiography in whole rodent brains. Governed by the competing processes of light attenuation through the brain tissues and optoacoustic signal generation due to absorption by blood, we identified three distinct spectral ranges centered around 580, 895 and 1100 nm optimally suited for imaging of vessels of different size and depth. The developed model was employed for numerical simulation of optoacoustic imaging of murine brain illustrating the effect of probing wavelength on visibility of the cerebral vasculature. In vivo imaging experiments further affirmed the capacity for whole-brain optoacoustic angiography in rodents at a wavelength of 1064nm.
A novel approach to monitor photosensitizer accumulation and photobleaching in the course of photodynamic therapy (PDT) with the use of nanoconstructs based on the simultaneous fluorescence (FL) and optoacoustic (OA) imaging is implemented. A liposome nanoconstructs employed in this studies contain benzoporphyrin derivatives (BPD) which serve as a photosensitizer and secondly, as a fluorophore, and the fluorescent IRDye800 dye acting as an additional contrasting agent due to its high quantum yield. FL provides visualization of BPD and IRDye800 distribution, while OA principle allows for BPD-absorption based imaging of tumor and its vascular environment. We demonstrate the results of a preliminary in vivo study with combined FL and OA custom-made setups on a NUDE mouse with human glioblastoma U- 87. The results of this studies show a hemorrhage in the tumor area on the OA images obtained @532 nm after PDT that is not visually detected, but confirmed with the following histological verification. Fast nanoconstructs accumulation (< 10 min) was observed using FL imaging with the concentration in tumor only 10% higher than in surrounding tissues. We believe that the ratio of nanoconstructs accumulation in tumor can be significantly increased using target approach.
We propose a new approach to monitoring of photodynamic therapy (PDT) of glioblastoma with the use of targeted nanoconstructs containing a photosensitizer (PS) benzoporphyrin derivative (BPD) and IRDye800 dye, antibodies for efficient accumulation of the drug in a tumor, and a chemotherapeutic agent for combined effect on tumor cells. Monitoring of PDT is based on the simultaneous fluorescent and optoacoustic (OA) imaging. Fluorescent imaging provides visualization of fluorescence agents with high molecular sensitivity, and monitoring of the effectiveness of PDT by PS photobleaching. OA allows to examine the vascular pattern of the tumor environment, as well as assess the tumor depth. IRDye800 is a better contrast agent in comparison to BPD due to red shifted spectral characteristics and higher fluorescence quantum yield. The results of numerical simulations have been verified in phantom studies using fluorescence and optoacoustic experimental setups and an agar phantom with optical characteristics similar to those of murine brain.
Raster-scan optoacoustic angiography at 532 nm wavelength with 50 μm lateral resolution at 2 mm diagnostic depth was used for quantitative characterization of neoangiogenesis in colon cancer models. Vessels of subcutaneously growing murine colon carcinoma (СT26) was imaged from 5th to 13th day of growth. The values of vascular density were calculated from the optoacoustic data. Inhomogeneous distribution of areas with high and low vascularization was demonstrated in the tumors. During tumor development vessel growth from the periphery to the center of the tumor was shown. Increase of vascular density precedes the increase of tumor volume. The obtained results may be important for the investigation of tumor development and for improvement of cancer treatment strategies.
KEYWORDS: In vivo imaging, In vitro testing, Blood, Absorption, Blood oxygen saturation, Signal attenuation, Optoacoustics, Blood vessels, Calibration, Veins
We present the comparison of two approaches of blood oxygen saturation determination from multispectral optoacoustic measurements: a calibration-free approach based on evaluation of the effective optical attenuation coefficient derived from in-depth OA signal decay, and an approach based on determination of optical absorption coefficient from OA signal amplitudes. Both approaches were tested in in vitro and in vivo experiments. The results of in vitro and in vivo experiments demonstrated the large difference between experimentally obtained μeff spectra and the literature data, that indicates much lower potential of the OA signal decay approach as compared to OA amplitudes approach. In vivo measurements of the μa spectrum experimentally obtained from OA signal amplitudes give the saturation values of 0.57±0.08 and 0.50±0.07 for two veins of the thoracic spine that agree well with physiological values for venous blood oxygenation in rat. Instead of multiple wavelengths measurements, a pair of wavelengths can be employed for OA measurements. In this case, the saturation maps were obtained at all wavelength pairs from the 658‒1069 nm range. The results demonstrated that the most accurate oxygenation values can be achieved at wavelength pairs of 700 nm and a wavelength from the range 850-1069 nm.
KEYWORDS: Sensors, Monte Carlo methods, Acoustics, In vivo imaging, Image resolution, Optical properties, Antennas, 3D image processing, 3D image reconstruction, Photoacoustic microscopy
We propose a hybrid approach to image enhancement in acoustic resolution photoacoustic microscopy. The developed technique is based on compensation for nonuniform spatial sensitivity of the optoacoustic (OA) system in both optical and acoustic domains. Spatial distribution of optical fluence is derived from full three-dimensional Monte Carlo simulations accounting for conical geometry of tissue laser illumination at the wavelength of 532 nm. Approximate nonuniform spatial response of acoustic detector with numerical aperture of 0.6 is derived from the two-dimensional k-Wave modeling. Application of the developed technique allows to improve the spatial resolution and to balance in-depth signal-level distribution in OA images of phantom and in-vivo objects.
We provide direct experimental comparison of the optoacoustic imaging performance of two different 64-element linear detector array (LDA) units based on polyvinylidene difluoride (PVDF) films. The first LDA unit was based on traditional flexible circuit (FC) technology and consisted of an FC glued to the nonmetalized signal surface of a 28-μm-thick PVDF film providing 300 / 80-μm axial resolution/lateral resolution (AR/LR) and 0.4-kPa noise equivalent pressure of its single element. The other LDA unit was manufactured using a technology of low-temperature photolithographic etching (PE) of a signal electrode onto a 25-μm-thick PVDF film providing 300 / 40-μm AR/LR and 1 kPa noise equivalent pressure. As compared with a previously reported LDA unit based on a 100-μm PVDF thick film, the main advantage of using the thinner PVDF films was 10-fold improvement in axial resolution, whereas the main drawback was 10-fold increased noise equivalent pressure. In terms of in vivo imaging performance, higher bandwidth of PE LDA probe was more important than the higher sensitivity of FC LDA unit.
The ability for noninvasive visualization of functional changes of a tumor’s oxygenation and circulatory system offers new advantages for prognosis and monitoring of the treatment efficacy. The results of breast cancer oxygen state study under chemotherapy action obtained by diffuse optical spectroscopy (DOS) in combination with Doppler ultrasonic imaging are presented. Complex use of optical and ultrasound methods gives complementary information about the size of the tumor node, peculiarities of its vascular bed, rate of its blood flow as well as oxygenation, and provide a picture of the tumor response to treatment. Comparison with tumor pathologic response allowed to identify differences in the changes of these parameters depending on the degree of pathological tumor response to chemotherapy. It was demonstrated that fourth and fifth degrees of therapeutic pathomorphism may be predicted by the increase of oxygen saturation level after the first cycle of chemotherapy. If the reduction or absence of the oxygen saturation dynamics is observed, first or second degree of pathological tumor response can be expected. Additional ultrasound investigation of the tumors may be useful for observation of the dynamics of tumor blood flow thereby for understanding the reasons of induced chemotherapy oxygenation changes. The proposed approach based on DOS and ultrasonography may be applied for monitoring of breast tumors under therapy and prediction of their sensitivity.
We will present reflection-mode bioimaging system providing complementary optical, photoacsoutic and acoustic measurements by acoustic detector after each laser pulse with 2kHz repetition rate. The photons absorbed within the biological tissue provide optoacoustic (OA) signals, the photons absorbed by the external electrode of a detector provide the measurable diffuse reflectance (DR) from the sample and the probing ultrasonic (US) pulse.
To demonstrate the in vivo capabilities of the system we performed complementary DR/OA/US imaging of small laboratory animals and human palm with 3.5mm/50μm/35μm lateral resolution at up to 3 mm diagnostic depth. Functional OA and DR imaging demonstrated the levels of tissue vascularization and blood supply. Structural US imaging was essential for understanding the position of vessels and zones with different perfusion.
Before BiOS-2017 we plan to accomplish more in vivo experiments validating the developed triple-modality system as diagnostic tool to detect vascularization as well as mechanisms of vascular changes when monitoring response to therapy.
Non-invasive measurement of blood oxygen saturation in blood vessels is a promising clinical application of optoacoustic imaging. However, unknown spatial and spectral distribution of optical fluence within biotissue challenges precise multispectral optoacoustic measurements of blood oxygen saturation. The accuracy of the blood oxygen saturation measurement can be improved by the choice of optimal laser wavelengths. We propose the numerical approach to determine the optimal wavelengths for two-wavelengths OA measurements of blood oxygen saturation at various depths. The developed approach accounts for acoustic pressure noise, error in determination of optical scattering and absorption coefficients used for the calculation of the optical fluence, and diameter of the investigated blood vessel. We demonstrate that in case of an unknown (or partially known) fluence spatial distribution at depths between 2 and 8 mm, minimal error in the determination of blood oxygen saturation is achieved at the wavelengths of 658±40 nm and 1069±40 nm. We report on the pilot results of OA in vivo measurements of blood oxygen saturation using optimal wavelengths obtained by the proposed approach.
Effective breast cancer treatment requires the assessment of metabolic changes of tumor tissue during chemo- and hormonotherapy for prediction tumor response. Evaluation of the dynamics of tumor oxygen state (by diffuse optical spectroscopy imaging) and tumor vasculature (by ultrasound investigation in power Doppler mode) was performed before treatment beginning and before the second cycle of chemotherapy in 16 patients who received preoperative chemotherapy. Changes of these indicators were compared then with tumor pathologic response. Breast tumors demonstrated different dynamics of tumor oxygenation depending on the changes of tumor tissue. The increase of the tumor oxygenation after the first cycle of chemotherapy was observed in five of six patients with grade 4 and 5 of pathologic tumor response. Decrease of the oxygenation level was revealed in one patient with the 4th degree of tumor response. Variable changes of the oxygenation level were mentioned in the patients with moderate (the 3d degree) tumor response. Tumor oxygenation decreased or was unchanged in case of 1 or 2 degree of tumor response in five of six cases. The study of the tumor blood vessels didn't reveal any correlation between vasculature changes and tumor response under the performed treatment. The trend of tumor oxygenation in early time after treatment beginning might be a predictive criterion of tumor sensitivity to chemotherapy.
The paper is dedicated to the analysis of applicability of passive acoustic brightness (PAB) method for noninvasive
detection and localization of optical heterogeneities under laser impact. To estimate the opportunities of PAB method
we used computer model of multilayered medium (similar to its optical and thermophysic properties to real human
tissue). It was demonstrated that PAB method allows to achieve high-quality diagnostics results in case of optical
heterogeneities located in depth range less than 20 millimeters. Topics of combining PAB method with Photo-Acoustic
(PAT) method were also discussed in terms of sensitizing of both approaches.
The opportunities and features of applying multi-focused antennae based on Fresnel lens to the problems of acoustical brightness thermometry have been studied. The technical feasibility of the device equipped with these antennae has been analyzed. The method has been suggested for the optimization of choosing radii of Fresnel lens rings providing acceptable focusing quality at various distances taking into account the peculiarities of acoustical brightness thermometry -- wide-band signal receiving in millimeter range and frequency-dependent acoustical absorption.
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