Photoacoustic imaging (PAI) provides optical contrast at depth beyond the optical transport mean free path. From the generation of ultrasound by light absorption, images can be reconstructed at the acoustic resolution ( 100 μm) with a penetration of a few cm. The design of imaging systems often leads to limited view artifacts, where a part of the information needed for a complete reconstruction of the objects is missing. We theoretically show that a dynamic approach based on the analysis of fluctuations induced by blood flow can suppress visibility artefacts. We demonstrate the performance of 3D Photoacoustic Fluctuation imaging (PAFI) using a spherical array with limited number of channels (256
elements, 8 MHz) in the chicken embryo model. Due to the low number of channels, standard PAI reconstructions additionally suffer from a poor contrast, which is enhanced by 2 to 3-fold using PAFI. We present an implementation of simultaneous PAFI and Ultrasound Power Doppler and present some results with coupled flow direction evaluations and optical contrast. Photoacoustic fluctuation imaging overcomes many limitations of conventional imaging and will be further evaluated for in-vivo imaging.
We investigate the use of a 256-channel spherically focused sparse array for volumic real-time ultrasound (US) and photoacoustic (PA) imaging of chicken embryo in vivo. Reconstructions were performed offline and signal processing techniques exploiting spatial and temporal dynamics of the blood flow were applied to visualize the vasculature. The resulting reduction of the clutter enhances the contrast by up to a factor of 2, providing an enhanced visualization of vascular networks.
This methodology has a potential for in vivo 3D real time visualization of the vasculature and other features using complementary information provided by US and PA imaging.
It has been shown that sub-diffraction structures can be resolved in acoustic resolution photoacoustic imaging thanks to norm-based iterative reconstruction algorithms exploiting prior knowledge of the point spread function (PSF) of the imaging system. Here, we demonstrate that super-resolution is still achievable when the receiving ultrasonic probe has much fewer elements than used conventionally (8 against 128). To this end, a proof-of-concept experiment was conducted. A microfluidic circuit containing five parallel microchannels (channel’s width 40μm, center-to center distance 180μm) filled with dye was exposed to 5ns laser pulses (=532nm, fluence=3.0mJ/cm2, PRF=100Hz). Photoacoustic signals generated by the sample were captured by a linear ultrasonic array (128 elements, pitch=0.1mm, fc=15MHz) connected to an acquisition device. The forward problem is modelled in a matrix form Y=AX, where Y are the measured photoacoustic signals and X is the object to reconstruct. The matrix A contained the PSFs at all points of the reconstruction grid, and was derived from a single PSF acquired experimentally for a 10-μm wide microchannel. For the reconstruction, we used a sparsity-based minimization algorithm. While the conventional image obtained by beamforming the signals measured with all the 128 elements of the probe cannot resolve the individual microchannels, our sparsity-based reconstruction leads to super-resolved images with only 8 elements of the probe (regularly spaced over the full probe aperture), with an image quality comparable to that obtained with all the 128 elements. These results pave the way towards super-resolution in 3D photoacoustic imaging with sparse transducers arrays.
Limited-view and bandwidth-limited acquisition in acoustic–resolution photoacoustic imaging lead to known imaging artifacts. To eliminate these artifacts, it has first been proposed to use fluctuations induced by multiple speckle illuminations to recover otherwise invisible features. However, a very small size of the optical speckle grain at depth in tissue against the acoustic resolution makes this approach unrealistic in practice. Here, we demonstrate experimentally in vitro that fluctuations induced by blood flow at physiological concentration may be exploited to improve visibility in photoacoustic imaging. We first illustrate how our method reveals features otherwise invisible due to the source directivity: a bended capillary tube (inner diameter 100µm) filled with blood flowing at 1.7cm/s was illuminated by 5ns laser pulses (=532nm, fluence=3.0mJ/cm2, PRF=100Hz) and imaged with a linear ultrasound array (128 elements, pitch=0.1mm, fc=15MHz) connected to an acquisition device. Being partially invisible in the conventional image, the whole capillary is reconstructed by means of a second-order analysis of photoacoustic images. Second, we illustrate how our approach allows for visualization of the inside of large objects otherwise invisible due to highpass filtering: we performed a second-order analysis on photoacoustic data resulting from illumination (=800nm, fluence=9.0mJ/cm2, PRF=10Hz/100Hz) of a glass tube (inner diameter 1mm) with blood flowing at 1cm/s. Whether the tube is perpendicular to the imaging plane or is lying inside the imaging plane parallel to the probe, the whole blood volume is visible on the fluctuation-based image, whereas conventional imaging only reveals the blood stream boundaries.
The STORM and PALM techniques developed in the past decade in optics allow resolving sub-wavelength structures based on localization. Here, we demonstrate that localization can be used to go beyond the diffraction limit in acoustic resolution photoacoustic imaging. To this end, a proof-of-concept photoacoustic localization experiment was conducted. A silicone sample containing five parallel microchannels (channel’s width is 40μm; center-to center distance equals 180μm), fed with a water suspension of 10μm red coated microbeads at a constant flow rate, was exposed to 5ns laser pulses (wavelength=532nm, fluence=3.0 mJ/cm2). At each laser pulse the microbeads produced a photoacoustic response that was then detected by a linear US array (128 elements, L22-8v, Verasonics, USA) connected to an acquisition device (High Frequency Vantage 256, Verasonics, USA). The design of the microfluidic circuit and the concentration of microbeads ensured sparse but random distribution of microbeads at any moment. The photoacoustic data was processed by a delay-and-sum algorithm, whose output was correlated with the system’s PSF function to obtain the position of microbeads at each laser shot. These positions were accumulated onto the localization grid providing a super-resolved image of the micro-fluidic circuit. Although being indistinguishable in a conventional US image, the microchannels dimensions and position were accurately reconstructed on the localization grid with 34.8±1.3μm for the channel’s width and 179±2.5μm for the center-to-center distance. As the first demonstration of super-localization in photoacoustics, these results constitute the first step towards imaging of red blood cells at depth beyond the acoustic diffraction limit.
The resolution of photoacoustic imaging of blood vasculature is limited at depth by the acoustic diffraction limit. In this work, we propose to exploit the fluctuations caused by flowing absorbers (such as red blood cells in blood vessels) to perform photoacoustic imaging beyond the acoustic diffraction limit: following the super-resolution optical fluctuation imaging (SOFI) method, we analyze the n-th order statistics from the temporal photoacoustic fluctuations induced by flowing particles.
We performed a proof-of-concept experiment in a 5-channel microfluidic silicon-based circuit flown with a suspension of RBC-mimicking 10 µm red-tainted polymer spheres (Microparticles, GmbH, Berlin, Germany). The sample was illuminated with a 5 ns pulsed ND-YAG laser (532 nm, Innolas, Krailling, Germany) with a fluence of 3 mJ/cm^2 and imaged at a 20 Hz rate using a L22-8v probe (128 elements, Verasonics, Redmond, WA, USA) coupled to a Verasonics Vantage 256 ultrasound scanner. Whereas the resolution of conventional photoacoustic imaging was too low to resolve individual channels, the nth order statistical analysis of the photoacoustic fluctuations provided images with a resolution enhancement scaling as n^{1/2}, in agreement with the SOFI theory and with numerical simulations. As opposed to our previous work which exploited speckle-based photoacoustic fluctuations to increase the resolution, the approach proposed here based on sample fluctuations do not require coherent light and can be readily applied to conventional photoacoustic imaging setup. Furthermore, in order to discard the oscillatory behavior of the photoacoustic point-spread-function, we extended in this work the SOFI theory to complex-valued photoacoustic images.
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