Axial Resolution Enhancement Research Articles

O-PRESS: Boosting OCT axial resolution with Prior guidance, Recurrence, and Equivariant Self-Supervision

Optical coherence tomography (OCT) is a noninvasive technology that enables real-time imaging of tissue microanatomies. The axial resolution of OCT is intrinsically constrained by the spectral bandwidth of the employed light source while maintaining a fixed center wavelength for a specific application. Physically extending this bandwidth faces strong limitations and requires a substantial cost. We present a novel computational approach, called as O-PRESS, for boosting the axial resolution of OCT with Prior guidance, a Recurrent mechanism, and Equivariant Self-Supervision. Diverging from conventional deconvolution methods that rely on physical models or data-driven techniques, our method seamlessly integrates OCT modeling and deep learning, enabling us to achieve real-time axial-resolution enhancement exclusively from measurements without a need for paired images. Our approach solves two primary tasks of resolution enhancement and noise reduction with one treatment. Both tasks are executed in a self-supervised manner, with equivariance imaging and free space priors guiding their respective processes. Experimental evaluations, encompassing both quantitative metrics and visual assessments, consistently verify the efficacy and superiority of our approach, which exhibits performance on par with fully supervised methods. Importantly, the robustness of our model is affirmed, showcasing its dual capability to enhance axial resolution while concurrently improving the signal-to-noise ratio.

Axial resolution enhancement of borehole acoustic measurements via inversion-based interpretation supported with ultrasonic data

Conventional borehole acoustic measurements deliver P and S wave slowness logs that inherently average in-situ rock properties along the receiver array of the acoustic instrument. These acquisition and processing conditions often limit the accuracy and resolution of the estimated rock elastic properties across heterolithic sedimentary sequences. We introduce an inversion-based interpretation method for borehole acoustic measurements that improves their vertical resolution by complementing them with ultrasonic borehole images. Results consist of high-resolution, layer-by-layer P and S wave slownesses. The combination of borehole acoustic measurements with borehole ultrasonic images enhances the definition of small rock features such as thin beds or vugs. We verify the new inversion-based interpretation method with synthetic borehole measurements and field acoustic logs acquired across sandstone-shale laminated formations and spatially heterogeneous carbonates. High-resolution layer-by-layer compressional and shear slownesses obtained with the new inversion method give rise to wider variations of calculated elastic properties than with standard acoustic logs for improved petrophysical and geomechanical evaluation. It is also found that implementing a common set of layers for the estimation of layer-by-layer rock elastic properties mitigates biases due to discrepancies in the intrinsic resolution of the various input measurements.

Broadband transparent ultrasound transducer with polymethyl methacrylate as matching layer for in vivo photoacoustic microscopy

Photoacoustic imaging (PAI) uniquely combines optics and ultrasound, presenting a promising role in biomedical imaging as a non-invasive and label-free imaging technology. As the traditional opaque ultrasound (US) transducers could hinder the transportation of the excitation light and limit the performance of PAI system, piezoelectric transparent ultrasonic transducers (TUTs) with indium tin oxide (ITO) electrodes have been developed to allow light transmission through the transducer and illuminate the sample directly. Nevertheless, without having transparent matching materials with appropriate properties, the bandwidth of those TUTs was generally narrow. In this work, we propose to employ polymethyl methacrylate (PMMA) as the matching layer material to improve the bandwidth of lithium niobate (LN)-based TUTs. The effects of PMMA matching layer on the performance of TUTs have been systematically studied. With the optimized PMMA matching layer, the very wide bandwidth of > 50 % could be achieved for the TUTs even with different transducer frequencies, leading to the great enhancement of axial resolution when compared to the similar reported work. In addition, the imaging performance of the developed TUT prototype has been evaluated in a PAI system and demonstrated by both phantom and in vivo small animal imaging.

Axial Resolution Enhancement of Optical Sectioning Structured Illumination Microscopy Based on Three-Beam Interference

As a branch of 3D microscopy, optical sectioning structured illumination microscopy (OS-SIM) has the advantages of fast imaging speed, weak photobleaching and phototoxicity, and flexible and compatible configuration. Although the method of using the one-dimensional periodic fringe pattern projected on the sample can remove the out-of-focus background from the in-focus signal, the axial resolution of the final reconstructed 3D image is not improved. Here, we propose a three-beam interference OS-SIM, namely TBOS, instead of the common-used dual-beam interference OS-SIM (DBOS). The three-beam interference scheme has been adopted in 3D super-resolution SIM (3D-SR-SIM), where the fringe phase shifting needs to be along each of the three orientations. In contrast, TBOS applies phase shifting only in one arbitrary direction. We built a TBOS SIM microscope and performed the 3D imaging experiments with 46 nm diameter fluorescent microspheres and a mouse kidney section. The axial resolution of the 3D image obtained with TBOS was enhanced by a factor of 1.36 compared to the DBOS method, consistent with the theoretical analysis and simulation. The OS-SIM with enhanced axial resolution for 3D imaging may find a wide range of applications in the biomedical field.

Axial resolution and imaging contrast enhancement in inverted light-sheet microscopy by natural illumination modulation.

Inverted light-sheet microscopy (ILSM) is widely employed for fast large-volume imaging of biological tissue. However, the scattering especially in an uncleared sample, and the divergent propagation of the illumination beam lead to a trade-off between axial resolution and imaging depth. Herein, we propose naturally modulated ILSM (NM-ILSM) as a technique to improve axial resolution while simultaneously maintaining the wide field-of-view (FOV), and enhancing imaging contrast via background suppression. Theoretical derivations, simulations, and experimental imaging demonstrate 15% axial resolution increases, and fivefold greater image contrast compared with conventional ILSM. Therefore, NM-ILSM allows convenient imaging quality improvement for uncleared tissue and could extend the biological application scope of ILSM.

Axial resolution enhancement for planar Airy beam light-sheet microscopy via the complementary beam subtraction method.

Airy beam light-sheet illumination can extend the field of view (FOV) of light-sheet fluorescence microscopy due to the unique propagation properties of non-diffraction and self-acceleration. However, the side lobes create undesirable out-of-focus background, leading to poor axial resolution and low image contrast. Here, we propose an Airy complementary beam subtraction (ACBS) method to improve the axial resolution while keeping the extended FOV. By scanning the optimized designed complementary beam that has two main lobes (TML), the generated complementary light-sheet has almost identical intensity distribution to that of the planar Airy light-sheet except for the central lobe. Subtraction of the two images acquired by double exposure respectively using the planar Airy light-sheet and the planar TML light-sheet can effectively suppress the influence of the out-of-focus background. The axial resolution improves from ∼4µm to 1.2 µm. The imaging performance was demonstrated by imaging specimens of aspergillus conidiophores and GFP labeled mouse brain section. The results show that the ACBS method enables the Airy beam light-sheet fluorescence microscopy to obtain better imaging quality.

Multiview tiling light sheet microscopy for 3D high-resolution live imaging.

Light-sheet or selective plane illumination microscopy (SPIM) is ideally suited for in toto imaging of living specimens at high temporal-spatial resolution. In SPIM, the light scattering that occurs during imaging of opaque specimens brings about limitations in terms of resolution and the imaging field of view. To ameliorate this shortcoming, the illumination beam can be engineered into a highly confined light sheet over a large field of view and multi-view imaging can be performed by applying multiple lenses combined with mechanical rotation of the sample. Here, we present a Multiview tiling SPIM (MT-SPIM) that combines the Multi-view SPIM (M-SPIM) with a confined, multi-tiled light sheet. The MT-SPIM provides high-resolution, robust and rotation-free imaging of living specimens. We applied the MT-SPIM to image nuclei and Myosin II from the cellular to subcellular spatial scale in early Drosophila embryogenesis. We show that the MT-SPIM improves the axial-resolution relative to the conventional M-SPIM by a factor of two. We further demonstrate that this axial resolution enhancement improves the automated segmentation of Myosin II distribution and of nuclear volumes and shapes.

Enhancement of axial resolution and image contrast of a confocal microscope by a microsphere working in noncontact mode.

A new technique, to the best of our knowledge, for improving the axial resolution and imaging contrast of a reflection mode confocal microscope is proposed. A 50 µm silica microsphere is added in front of the objective lens to enhance both the focusing of illumination and the collection of reflected and scattered light from sample surfaces in noncontact mode. An adjustable pinhole is used to compensate the displacement of the focal point in the axial direction. Various samples, including grouped nanolines and nanosteps, are used to demonstrate imaging performance. By comparison to an NA 0.9 commercial confocal microscope, the new setup achieves the axial resolution up to 100 nm and increases the image contrast by 4.56 times. The entire setup offers a cost-effective solution for high imaging performance, which can be applied in many fields from nanotechnology to biology.

Truncated correlation photoacoustic coherence tomography: An axial resolution enhancement imaging modality

In this report we present a novel photoacoustic (PA) modality using pulsed chirp excitation at a fixed wavelength and spectral analysis based on frequency-domain (FD) processing. We introduce Truncated Correlation Photoacoustic Coherence Tomography (TC-PACT), a PA axial resolution enhancement methodology, with an application to closely stacked subsurface multilayers of plexiglass coated with a thin layer of graphite acting as surface absorber. The experimental results demonstrated that the SNR and the axial resolution were higher in TC-PACT than in conventional FD photoacoustics. A ns pulsed laser was also used as PA excitation source with axial resolution results found to be commensurate with TC-PACT. However, the separation distance between adjacent absorbers when the latter approached the axial resolution limit of the transducer was only resolved in TC-PACT compared to pulsed laser PA owing to the lower uncertainty in the former modality involved in determining the location of the absorbers.

In-vivo sub-diffraction adaptive optics imaging of photoreceptors in the human eye with annular pupil illumination and sub-Airy detection.

Adaptive optics scanning light ophthalmoscopy (AOSLO) allows non-invasive visualization of the living human eye at the microscopic scale; but even with correction of the ocular wavefront aberrations over a large pupil, the smallest cells in the photoreceptor mosaic cannot always be resolved. Here, we synergistically combine annular pupil illumination with sub-Airy disk confocal detection to demonstrate a 33% improvement in transverse resolution (from 2.36 to 1.58 μm) and a 13% axial resolution enhancement (from 37 to 32 μm), an important step towards the study of the complete photoreceptor mosaic in heath and disease. Interestingly, annular pupil illumination also enhanced the visualization of the photoreceptor mosaic in non-confocal detection schemes such as split detection AOSLO, providing a strategy for enhanced multimodal imaging of the cone and rod photoreceptor mosaic.

Axial resolution analysis in compressive digital holographic microscopy.

Digital holographic microscopy with compressive sensing (CDHM) has successfully achieved tomography and has been applied in many fields. However, the enhancement of axial resolution in CDHM remains to be elucidated. By deducing accurate formulas for the lateral and axial resolutions without paraxial approximation, we quantized the elongation effect of a digital holography (DH) system in this study. Thus, we revealed that the elongation effect, which is affected only by the system's numerical aperture (NA), is an inherent property of DH systems. We present a detailed analysis herein on the physical significance of the coherence parameter, which is the ratio of a system's limit axial resolution to the interlayer spacing more thoroughly than in previous research. Further, we achieved the tomography of a fiber by using a DH system with a 10 × microscope, with CS to eliminate the elongation effect, and experimentally validated our theoretical results. By applying these theoretical guidelines, we distinguished crossed fibers at distances of 36.4 μm and 48.5 μm, respectively, using the same experimental setup. There would be potential applications of this theory in tomography and observation of microscale objects in the areas of biological and fluid.

Cross-sectional imaging through scattering media by quantum-mimetic optical coherence tomography with wavefront shaping

Quantum-mimetic (QM) optical coherence tomography (OCT) combined with wavefront shaping is proposed as a novel approach to achieve high-resolution cross-sectional imaging through a strongly scattering medium. Specifically, QM-OCT is a recently devised OCT technique mimicking quantum OCT to acquire its main characteristic features, most notably immunity to group-velocity dispersion and enhancement of axial resolution, without quantum light. To validate the proposed method, we demonstrate experimentally that a one-dimensional cross-sectional image of a sample located behind a diffuser plate is obtained successfully and efficiently with QM-OCT by shaping the wavefront of light incident onto the diffuser.

3D resolution enhancement in saturated competition microscopy

To overcome the diffraction barrier, super-resolution microscopy is contrived and has witnessed scientific developments in varying fields, especially in last few decades, such as stochastic optical reconstruction microscopy, stimulated emission depletion microscopy (STED), mirror-enhanced super-resolution microscopy (MEANS), and fluorescence emission difference microscopy (FED). Recently, saturated competition microscopy (SAC) was developed to realize high sub-diffraction resolution in either fluorescent or non-fluorescent imaging. Compared with STED, SAC features non-constraint in fluorescent dye selection. Nevertheless, the lateral resolution is limited in consideration of photobleaching side effects. Also, the axial resolution enhancement of SAC has not been demonstrated. In this study, a method, combining FED, MEANS, and SAC, is presented to improve the three-dimensional (3D) resolution. The numerical study reveals that the lateral resolution is close to 0.085λ and axial resolution can be enhanced to 0.184λ. In addition, the SNR is improved simultaneously. The availability to improve 3D resolution of SAC is believed to be significant for biological imaging in the future.

Axial resolution and contrast enhancement in digital scanned light-sheet microscopy via stimulated emission depletion

Integration of stimulated emission depletion (STED) with digital scanned light-sheet microscopy (DSLM) using a Bessel beam achieves fast, high-resolution and large field of view (FOV) imaging. Here, we report on a combination method of DSLM and STED by using a Bessel beam for excitation and a hollow Gaussian beam for depletion. Owing to the advantages of confined axial distribution around the focal plane for the hollow Gaussian depletion beam, those fluorescence signals from the periphery of the Bessel excitation pattern are depleted by stimulated emission. A super-resolved light sheet with suppressed side lobes is achieved. The numerical simulation results show that the use of the Bessel beam for the excitation and the hollow Gaussian beam for the STED results in a DSLM system with enhanced performance of the axial resolution and image contrast over a moderately large FOV.

Spectroscopic optical coherence refraction tomography.

In optical coherence tomography (OCT), the axial resolution is often superior to the lateral resolution, which is sacrificed for long imaging depths. To address this anisotropy, we previously developed optical coherence refraction tomography (OCRT), which uses images from multiple angles to computationally reconstruct an image with isotropic resolution, given by the OCT axial resolution. On the other hand, spectroscopic OCT (SOCT), an extension of OCT, trades axial resolution for spectral resolution and hence often has superior lateral resolution. Here, we present spectroscopic OCRT (SOCRT), which uses SOCT images from multiple angles to reconstruct a spectroscopic image with isotropic spatial resolution limited by the OCT lateral resolution. We experimentally show that SOCRT can estimate bead size based on Mie theory at simultaneously high spectral and isotropic spatial resolution. We also applied SOCRT to a biological sample, achieving axial resolution enhancement limited by the lateral resolution.

Enhancing the axial resolution of two-photon imaging.

An axial-resolution-enhanced two-photon laser scanning microscopy system is presented in this paper. In the proposed method, we use a spatial light modulator (SLM) for the phase modulation of the excitation light. The axially split point spread function (PSF) is generated by loading a 0-π pattern on the SLM. The final quality-enhanced images are acquired by subtracting the two consecutive images acquired by the original PSF and the split PSF. Because of the fluorescence differential processing, the axial elongation of the particles images is suppressed, and the axial resolution is enhanced accordingly. With the axial resolution enhanced, the overlap between layer images is also reduced, which decreases the background noise of the images and enhances the contrast and image quality of the acquired fluorescence images. The capability of axial resolution and contrast enhancement is successfully demonstrated by both theoretical calculation and experimental results.

Sidelobe suppression and axial resolution enhancement in 4pi microscopy with higher-order radially polarized Laguerre–Gaussian beams using subtractive imaging

Subtractive imaging is used to suppress the axial sidelobes and improve the axial resolution of 4pi microscopy with a higher-order radially polarized (RP) Laguerre–Gaussian (LG) beam. A solid-shaped point spread function (PSF) and a doughnut-shaped PSF with a dark spot along the optical axis are generated by tightly focusing a higher-order RP-LG beam and a modulated circularly polarized beam, respectively. By subtracting the two images obtained with those two different PSFs, the axial sidelobes of the subtracted PSF are reduced from 37% to about 10% of the main lobe, and the axial resolution is increased from 0.21λ to 0.15λ.

Supercritical focusing coherent anti-Stokes Raman scattering microscopy for high-resolution vibrational imaging.

We report the development of, to the best of our knowledge, a novel supercritical focusing coherent anti-Stokes Raman scattering (SCF-CARS) microscopy for high-resolution vibrational imaging. Two optimized phase patterns with a combination of concentric rings with an alternative 0 and π phase are generated by using a spatial light modulator and applied to the pump beam for minimizing its focal spot size. One of the phase patterns is for both the lateral and axial resolution enhancement, and the other can further improve the lateral resolution, but it sacrifices the axial resolution to some extent. We demonstrate this high-resolution SCF-CARS microscopy technique by imaging the polymethyl methacrylate (PMMA) nano-cylinder on a microscope slide and glass-air interface, as well as biomedical samples, for example, tooth.

Axial resolution enhancement of light-sheet microscopy by double scanning of Bessel beam and its complementary beam.

The side lobes of Bessel beam will create significant out-of-focus background when scanned in light-sheet fluorescence microscopy (LSFM), limiting the axial resolution of the imaging system. Here, we propose to overcome this issue by scanning the sample twice with zeroth-order Bessel beam and another type of propagation-invariant beam, complementary to the zeroth-order Bessel beam, which greatly reduces the out-of-focus background created in the first scan. The axial resolution can be improved from 1.68 μm of the Bessel light-sheet to 1.07 μm by subtraction of the two scanned images across a whole field-of-view of up to 300 μm × 200 μm × 200 μm. The optimization procedure to create the complementary beam is described in detail and it is experimentally generated with a spatial light modulator. The imaging performance is validated experimentally with fluorescent beads as well as eGFP-labeled mouse brain neurons.

Breaking the Axial Diffraction Limit: A Guide to Axial Super‐Resolution Fluorescence Microscopy

AbstractOptical microscopy is a powerful tool for understanding the fundamentals of the microscopic world. However, for centuries its resolving ability remained limited by the optical diffraction limit. Super‐resolution fluorescence microscopy (SRFM) has been introduced to break the diffraction limit and significantly expand the fields in which optical microscopy can be applied. Unfortunately, SRFM contributes little towards axial resolution enhancement, rendering observation of the axial and three‐dimensional structures of biological tissues difficult; this may yield a misunderstanding of intracellular interactions. Based on the existing literature, the development of axial SRFM is still behind that of lateral SRFM. In light of this, this Review presents a comprehensive summary of the principles, development, characteristics, and applications of existing techniques for improving the axial resolution. This Review will provide a guide to researchers and promote further development of related technology.