Comparison of MEMS-based photoacoustic microscopy in biomedical imaging

Photoacoustic microscopy (PAM) is a biological visualization technique that can provide high spatial resolution and high contrast images of deep structures in living tissues. However, because of the spherical aberration of the objective lens and the wavefront distortion due to the surface shape and light scattering of the specimen, obtained photoacoustic images in deep tissues are sometimes blurred or distorted. In order to solve this problem, we have developed a PAM using a transmissive liquid-crystal adaptive optics (AO) element. The transmissive and thin structure of the AO element can be easily installed in the PAM system. Using photoacoustic images of a USAF 1951 resolution test target measured through the glass substrate (thickness; 1.5-mm), the lateral resolutions in PAM were estimated with and without the AO element, when a flashlamp-pumped nanosecond pulse laser (pulse width, 5-ns; wavelength, 500-nm) and water-immersion objective lens (NA = 0.8) were employed. The lateral resolution of PAM at the depth of 1.5-mm was improved from 1.04 ± 0.04 μm to 0.53 ± 0.10 μm by optimizing AO corrections. We have also visualized small blood vessels in mouse ear in vivo by PAM with AO correction. Thus, by optimizing the AO correction according to the imaging depth, our proposed PAM improves the spatial resolution in biological tissues.

Compressive Sampling Photoacoustic Microscope System based on Low Rank Matrix Completion

ADMM based low-rank and sparse matrix recovery method for sparse photoacoustic microscopy

Photoacoustic microscopy (PAM) is a biological visualization technique that can provide high spatial resolution and high contrast images of deep structures in living tissues. In PAM, the lateral resolution is determined by the size of the focus spot. Generally, because the wavefront aberration, due to the difference of refractive index between samples and air (water) and the shape of samples, enlarges the focus spot, obtained deep images are blurred or distorted. In order to solve this problem, we corrected the wavefront aberration occurring in samples using a transmissive liquid-crystal adaptive optics (AO) element. Our AO element consists of three liquid-crystal layers which have different ITO (indium tin oxide) patterns and are controlled independently. Their patterns are designed to correct the wavefront aberration suitable for a 40X waterimmersion objective lens. The AO element with transmissive and thin structure is easily installed in the PAM system. Also, our AO element is inexpensive and has low power consumption. In this study, we compared photoacoustic images obtained without and with the AO element for a USAF test target, polystyrene beads diffused in glycerol and various tissue specimens. As a result, we found that the use of transmissive AO element improves the lateral resolution and signal-tonoise ratio in PAM.

As two important acoustic imaging modalities, photoacoustic microscopy (PAM) and ultrasound microscopy (UM) provide complementary functional and structural information on biological tissues. At penetration depths beyond the optical diffraction limit, the spatial resolution of both PAM and UM is determined acoustically by the receiving (focused) ultrasound transducer. To obtain good dual-modal imaging resolution, the transducer should have a high center frequency with a wide bandwidth. More importantly, a high acoustic numerical aperture (NA) should be maintained for receiving the PA and ultrasound signals, which is however difficult to achieve as the light blockage by the transducer is more likely to occur at higher NAs. In this paper, we report the dual-modal PAM and UM based on an optically-transparent focused PVDF transducer with a high NA of 0.64. The transducer has an acoustic center frequency and bandwidth of 36 MHz and 44 MHz, respectively. The acoustic focal diameter and zone are 37.8 μm and 210 μm, respectively. With a central transparent window, the excitation laser pulses can directly pass through the transducer to illuminate the target. This allows a high acoustic NA to be obtained without light blockage. For demonstration, co-registered 3D and 2D PAM/UM imaging experiments have been conducted on a twisted wire target in both water and chicken breast tissue, and in-vivo on a mouse tail, respectively. The imaging results show that high acoustic resolution and sensitivity can be achieved to resolve the target at different depths with a simple and compact dual-modal imaging setup

Photoacoustic (PA) imaging is one of the fastest growing imaging technologies nowadays in both research and clinical applications, especially due to its unique capability to visualize blood vessels. The PA microscopy (PAM) is classified into two types: optical-resolution PAM (OR-PAM) and acoustic-resolution PAM (AR-PAM). OR-PAM image has a point spread function (PSF) much smaller than AR-PAM because it uses a tightly focused optical beam and the PSF is determined by the optical focus. In contrast, AR-PAM uses an unfocused optical illumination to excite a relatively large area and detects the PA signal from a small area determined by its acoustic focus. Because ultrasound is less scattered than light in biological tissue, AR-PAM can achieve deeper imaging depth than OR-PAM at the expense of image resolution. Due to the limited resolution and imaging depth scale of each PAM type, it is challenging to image vessels in various area of small animals. In this study, we demonstrated in vivo OR-/AR-PAM imaging of blood vessels in various areas such as eye, ear, and hind limb by using a single commercial PAM system. Additionally, we quantified micro-vessel density (MVD) of the mouse eye and ear images, and applied a synthetic aperture focusing technique (SAFT) to correct the distorted PA signal at the out-of-focus in AR-PAM image. As a result, we have demonstrated multiscale PAM imaging of small animal vasculature in various areas with vessel quantification and resolution enhancement, so we believe that this multiscale PAM imaging technique would be helpful in biology research such as ischemia and neovascularization.

Photoacoustic microscopy (PAM) is a high-resolution imaging modality capable of visualizing fine microvasculature in a biological tissue. Clinical translation of PAM system is still an issue due to the use of expensive and high energy laser sources for imaging. Although low energy laser sources can facilitate clinical transition of PAM systems as they are rugged, portable, affordable, and safe to use, the photoacoustic (PA) signal they generate is very weak, resulting in very low signal-to-noise ratio (SNR) PA signals and in turn low quality PA images. In this study, we have developed an enhancement autoencoder (EAE) utilizing fully convolutional neural network, that can improve the quality of the PA signals received, consequently improving the SNR of the reconstructed images. We acquired PAM data from rat brain tissue with both high energy (target data) and low energy (input data) of the laser for training purposes and tested our trained model on PAM data obtained from new rat brains. Our effort is to reconstruct the vascular structure as well as an accurate reading for the blood concentration. The latter has been neglected in the previous studies. The performance of our EAE is evaluated in terms of SNR, structural similarity index (SSIM), root mean square error (RMSE) and correlation.

As a high-resolution deep tissue imaging technology, photoacoustic microscopy (PAM) is attracting extensive attention in biomedical studies. PAM has trouble in achieving real-time imaging with the long data acquisition time caused by point-to-point sample mode. In this paper, we propose a sparse photoacoustic microscopy reconstruction method based on matrix nuclear norm minimization. We use random sparse sampling instead of traditional full sampling and regard the sparse PAM reconstruction problem as a nuclear norm minimization problem, which is efficiently solved under alternating direction method of multiplier (ADMM) framework. Results from PAM experiments indicate the proposed method could work well in fast imaging. The proposed method is also be expected to promote the achievement of PAM real-time imaging.

High-resolution photoacoustic microscopy with deep penetration through learning

It is always a great challenge for pure optical techniques to maintain good resolution and imaging depth at the same time. Photoacoustic imaging is an emerging technique which can overcome the limitation by pulsed light illumination and acoustic detection. Here, we report a Near Infrared Acoustic-Resolution Photoacoustic Microscopy (NIR-AR-PAM) systm with 30 MHz transducer and 1064 nm illumination which can achieve a lateral resolution of around 88 μm and imaging depth of 9.2 mm. Compared to visible light NIR beam can penetrate deeper in biological tissue due to weaker optical attenuation. In this work, we also demonstrated the in vivo imaging capabilty of NIRARPAM by near infrared detection of SLN with black ink as exogenous photoacoustic contrast agent in a rodent model.

Photoacoustic microscopy with high spatial resolution and fast imaging acquisition allows observing dynamic processes of optical absorption-based microanatomic structures in three dimensions. An evanescent sensor accesses ultrasonic detection with high sensitivity and broad bandwidth while suffering from limited field of view (FOV), thus compromising the photoacoustic imaging acquisition rate. Here, we develop an optical-scanning evanescent sensor by fast deflection of the interrogation light along the interface of prism and water using a one-dimensional galvanometer, demonstrating excellent detection sensitivity of ∼132 Pa with a broadband frequency response of >140-MHz at an enlarged FOV of ∼2.90 × 0.19 mm2. Incorporating the optical-scanning evanescent sensor in photoacoustic microscopy, a volumetric image (∼3.0 × 0.25 × 1.0 mm3) with micrometer-scale spatial resolution is acquired within ∼2.5 s by synergistically scanning both photoacoustic illumination laser and sensor's interrogation light. High-speed imaging of flowing microparticles within a capillary tube offers the visualizations of the traveling processes in three dimensions. Potentially, the optical-scanning evanescent sensor allows photoacoustic microscopy accommodating to dynamic imaging at cellular level such as in vivo flow cytometry of circulating tumor cells.

Small animal models, such as zebrafish, drosophila, C. elegan, is considered to be important models in comparative biology and diseases researches. Traditional imaging methods primarily employ several optical microscopic imaging modalities that rely on fluorescence labeling, which may have potential to affect the natural physiological progress. Thus a label-free imaging method is desired. Photoacoustic (PA) microscopy (PAM) is an emerging biomedical imaging method that combines optical contrast with ultrasonic detection, which is highly sensitive to the optical absorption contrast of living tissues, such as pigments, the vasculature and other optically absorbing organs. In this work, we reported the whole body label-free imaging of zebrafish larvae and drosophila pupa by PAM. Based on intrinsic optical absorption contrast, high resolution images of pigments, microvasculature and several other major organs have been obtained in vivo and non-invasively, and compared with their optical counterparts. We demonstrated that PAM has the potential to be a powerful non-invasive imaging method for studying larvae and pupa of various animal models.

As the body's largest solid organ, the liver plays a very important role in the body's metabolism. More and more studies have shown that the liver may be an immune organ. A large amount of blood containing nutrients and pathogenic bacteria enters the liver. The liver is extremely rich in blood vessels, including the hepatic artery and portal vein. In the liver circulation, hepatic sinusoids have a unique structure and maintain normal immune homeostasis. More importantly, many liver diseases will directly affect liver sinusoids, such as hepatitis and liver fibrosis. Therefore, the ability to obtain high-resolution images of liver sinusoids of living livers is of great significance for the study of liver diseases. Photoacoustic imaging is becoming a very promising tool for the research of living organisms. It combines the high contrast of optical imaging and the high resolution of acoustic imaging to realize the imaging of absorption clusters in biological tissues. Since the scattering of ultrasound signals in biological tissues is 2-3 orders of magnitude weaker than the scattering of light in biological tissues, the endogenous absorption difference of tissues is directly used in the imaging process, so photoacoustic imaging has the advantages of deep imaging depth and non-destructive. As an important branch of photoacoustic imaging, photoacoustic microscopy can provide micron-level or even sub-micron-level imaging resolution, which is of great significance for biological research such as liver blood vessel detection. Since the lateral resolution of the photoacoustic microscopy imaging system depends on the focus of the laser, a higher resolution can be obtained by increasing the numerical aperture of the condenser objective. However, a large numerical aperture usually means a shorter working distance and makes the entire imaging system very sensitive to small optical defects. Therefore, the improvement of resolution through this method will be limited in practical applications. This paper implements a method of using iterative deconvolution to obtain a high-resolution photoacoustic image of the liver blood vessel. The focal spot of the photoacoustic microscopy is measured to obtain the lateral PSF (point spread function) of the system. Making the measured PSF as the initial system PSF to perform Lucy- Richardson (LR) deconvolution. The image resolution of liver lobules obtained by this method is higher. The full width at half maximum (FWHM) of the same liver sinusoid width before and after deconvolution are 9.52 μm and 6.65 μm, respectively, and the image definition is increased by about 1.4 times. Experiments show that this method can further improve the clarity of photoacoustic images of liver blood vessels, which lays the foundation for further research on liver diseases.

Photoacoustic imaging is a functional imaging method based on the photoacoustic effect, which combines the high contrast of optical imaging and the low dispersion characteristics of acoustic imaging. It has been developed rapidly in recent years. Photoacoustic microscopy, as an important branch, is widely used in biomedical imaging due to its high resolution, high contrast, and non-destructive characteristics. Compared with other medical imaging techniques, photoacoustic microscopy is simple, effective, low-cost, and does not generate ionizing radiation. But due to the need to focus the laser strongly, the depth of field is limited. Currently, most photoacoustic microscopy adopts an array scanning mechanism. Limited by the imaging depth of field and inherent scanning methods, photoacoustic microscopy cannot achieve large-volume, highresolution, and high-speed imaging at the same time. High-speed and large-scale tissue imaging is of great significance for the study of response mechanisms and the development of physiological and pathological processes. It is conducive to make accurate judgments for disease diagnosis. For the problems of photoacoustic microscopy, this paper proposes a highresolution three-dimensional information fusion technology suitable for photoacoustic microscopy, which provides richer structural and functional information for related physiological and pathological research. Combining the tomographic features of the data collected by the photoacoustic microscopy system with the traditional Laplacian pyramid transform, the depth of field of the photoacoustic microscopy is expanded. Two sets of single-focus image data sets of virtual photoacoustic microscopy platforms are collected and processed by three-dimensional high-resolution information fusion technology. The results demonstrates that a complete large-scale three-dimensional high-resolution structure has been successfully realized, and the reliability of the method is verified.

Photoacoustic imaging is a promising technique that combines optical contrast with ultrasonic detection to map the distribution of the absorbing pigments in biological tissues. It has been widely used in biological researches, such as structural imaging of vasculature, brain structural and functional imaging, and tumor detection. Photoacoustic microscopy (PAM) is an important branch of Photoacoustic imaging that can achieve spatial from the micron to submicron level. However, Common photoacoustic microscopy can only be fixed on a single scale (single resolution) for imaging, which makes it difficult to obtain rich information of biological tissue, and cannot adapt to different imaging needs. Here, we developed multi-scale photoacoustic microscopy by integrating optical-resolution photoacoustic microscopy and the acoustic-resolution photoacoustic microscopy using multi-dimension optical fiber. This method mainly uses a single mode fiber and a multimode fiber with same length to split a single laser pulse into two sub-pulses. Finally, the two laser pulses are focused on the sample by the objective lens, which can generate a focal spot covering a few microns to tens of microns near the focal plane. The size of the focal spot is determined by the core diameter of the fiber, and the different size of the spot leads to different lateral resolution, Therefore, the system can achieved both optical resolution (4 μm) and acoustic resolution (40 μm). The multi-scale imaging performance of the system is verified by imaging the blood vessels of mouse and cerebral vascular. The multiscale imaging capability of our system may fulfill different requirements in biomedical researches and facilitate multiscale interpretation in system biology.