AI-guided histopathology predicts brain metastasis in lung cancer patients.

Brain metastases can occur in nearly half of patients with early and locally advanced (stage I-III) non-small cell lung cancer (NSCLC). There are no reliable histopathologic or molecular means to identify those who are likely to develop brain metastases. We sought to determine if deep learning (DL) could be applied to routine H&E-stained primary tumor tissue sections from stage I-III NSCLC patients to predict the development of brain metastasis. Diagnostic slides from 158 patients with stage I-III NSCLC followed for at least 5 years for the development of brain metastases (Met+, 65 patients) versus no progression (Met-, 93 patients) were subjected to whole-slide imaging. Three separate iterations were performed by first selecting 118 cases (45 Met+, 73 Met-) to train and validate the DL algorithm, while 40 separate cases (20 Met+, 20 Met-) were used as the test set. The DL algorithm results were compared to a blinded review by four expert pathologists. The DL-based algorithm was able to distinguish the eventual development of brain metastases with an accuracy of 87% (p < 0.0001) compared with an average of 57.3% by the four pathologists and appears to be particularly useful in predicting brain metastases in stage I patients. The DL algorithm appears to focus on a complex set of histologic features. DL-based algorithms using routine H&E-stained slides may identify patients who are likely to develop brain metastases from those who will remain disease free over extended (>5 year) follow-up and may thus be spared systemic therapy. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Interferometric speckle visibility spectroscopy (iSVS) for measuring decorrelation time and dynamics of moving samples with enhanced signal-to-noise ratio and relaxed reference requirements.

Diffusing wave spectroscopy (DWS) is a group of techniques used to measure the dynamics of a scattering medium in a non-invasive manner. DWS methods rely on detecting the speckle light field from the moving scattering medium and measuring the speckle decorrelation time to quantify the scattering medium's dynamics. For DWS, the signal-to-noise (SNR) is determined by the ratio between measured decorrelation time to the standard error of the measurement. This SNR is often low in certain applications because of high noise variances and low signal intensity, especially in biological applications with restricted exposure and emission levels. To address this photon-limited signal-to-noise ratio problem, we investigated, theoretically and experimentally, the SNR of an interferometric speckle visibility spectroscopy (iSVS) compared to more traditional DWS methods. We found that iSVS can provide excellent SNR performance through its ability to overcome camera noise. We also proved an iSVS system has more relaxed constraints on the reference beam properties. For an iSVS system to function properly, we only require the reference beam to exhibit local temporal stability, while incident angle, reference phase and intensity uniformity do not need to be constrained. This flexibility can potentially enable more unconventional iSVS implementation schemes.

Robust Kramers-Kronig holographic imaging with Hilbert-Huang transform.

Holography based on Kramers-Kronig relations (KKR) is a promising technique due to its high-space-bandwidth product. However, the absence of an iterative process limits its noise robustness, primarily stemming from the lack of a regularization constraint. This Letter reports a generalized framework aimed at enhancing the noise robustness of KKR holography. Our proposal involves employing the Hilbert-Huang transform to connect the real and imaginary parts of an analytic function. The real part is initially processed by bidimensional empirical mode decomposition into a series of intrinsic mode functions (IMFs) and a residual term. They are then selected to remove the noise and bias terms. Finally, the imaginary part can be obtained using the Hilbert transform. In this way, we efficiently suppress the noise in the synthetic complex function, facilitating high-fidelity wavefront reconstruction using ∼20% of the exposure time required by existing methods. Our work is expected to expand the applications of KKR holography, particularly in low phototoxicity biological imaging and other related scenarios.

Implementation of free-space Fourier Ptychography with near maximum system numerical aperture.

Over the past decade, the research field of Fourier Ptychographic Microscopy (FPM) has seen numerous innovative developments that significantly expands its utility. Here, we report a high numerical aperture (NA) FPM implementation that incorporates some of these innovations to achieve a synthetic NA of 1.9 - close to the maximum possible synthetic NA of 2 for a free space FPM system. At this high synthetic NA, we experimentally found that it is vital to homogenize the illumination field in order to achieve the best resolution. Our FPM implementation has a full pitch resolution of 266 nm for 465 nm light, and depth of field of 3.6 µm. In comparison, a standard transmission microscope (incoherent) with close to maximum possible NA of 0.95 has a full pitch resolution of 318 nm for 465 nm light, and depth of field of 0.65 µm. While it is generally assumed that a free-space coherent imaging system and a free-space incoherent imaging system operating at their respective maximum NA should give comparable resolution, we experimentally find that an FPM system significantly outperforms its incoherent standard microscopy counterpart in resolution by a factor of 20%. Coupled with FPM's substantially longer effective depth of field (5.5 times longer), our work indicates that, in the near-maximum NA operation regime, the FPM has significant resolution and depth of field advantages over incoherent standard microscopy.

Imaging through highly scattering human skulls with ultrasound-modulated optical tomography.

Advances in human brain imaging technologies are critical to understanding how the brain works and the diagnosis of brain disorders. Existing technologies have different drawbacks, and the human skull poses a great challenge for pure optical and ultrasound imaging technologies. Here we demonstrate the feasibility of using ultrasound-modulated optical tomography, a hybrid technology that combines both light and sound, to image through human skulls. Single-shot off-axis holography was used to measure the field of the ultrasonically tagged light. This Letter paves the way for imaging the brain noninvasively through the skull, with optical contrast and a higher spatial resolution than that of diffuse optical tomography.

Single-shot surface 3D imaging by optical coherence factor.

We report a single-shot three-dimensional (3D) topographical imaging method, optical coherence factor (OCF) imaging, which uses optical coherence as the contrast mechanism to acquire the surface height (${z}$z-direction) information of an object. A 4-f imaging system records the light field reflected from the surface of the object. The illumination of the imaging system comes from a laser source with the optical coherence length comparable to the depth of field (DoF) of the optical system. Off-axis holographic recording is used to retrieve the coherence factor from the interference fringes, which is then converted to ${z}$z-direction information. In this experiment, we validate our 3D imaging results comparing them to axial scanning full-field optical coherence tomography images. We also analyze the contrast mechanism of OCF and show that it is able to provide additional information over conventional coherent and incoherent imaging using the same imaging setup. This single-shot computationally efficient method may have potential applications in industrial quality control inspection.

Computational aberration correction of VIS-NIR multispectral imaging microscopy based on Fourier ptychography.

Due to the chromatic dispersion properties inherent in all optical materials, even the best-designed multispectral objective will exhibit residual chromatic aberration. Here, we demonstrate a multispectral microscope with a computational scheme based on the Fourier ptychographic microscopy (FPM) to correct these effects in order to render undistorted, in-focus images. The microscope consists of 4 spectral channels ranging from 405 nm to 1552 nm. After the computational aberration correction, it can achieve isotropic resolution enhancement as verified with the Siemens star sample. We image a flip-chip to show the promise of our system to conduct fault detection on silicon chips. This computational approach provides a cost-efficient strategy for high quality multispectral imaging over a broad spectral range.

Wide-angular-range and high-resolution beam steering by a metasurface-coupled phased array.

Optical beam steering has broad applications in lidar, optical communications, optical interconnects, and spatially resolved optical sensors. For high-speed applications, phased-array-based beam-steering methods are favored over mechanical methods, as they are unconstrained by inertia and can inherently operate at a higher speed. However, phased-array systems exhibit a tradeoff between angular range and beam divergence, making it difficult to achieve both a large steering angle and a narrow beam divergence. Here, we present a beam-steering method based on wavefront shaping through a disorder-engineered metasurface that circumvents this range-resolution tradeoff. We experimentally demonstrate that, through this technique, one can continuously steer an optical beam within a range of 160° (80° from normal incidence) with an angular resolution of about 0.01° at the cost of beam throughput.

3D imaging scanner.

This paper proposes a multi-view three-dimensional display method based on a scanning imaging system with the light-intensity characteristic recorded by an improved flatbed scanner. Within the effective scanning depth of the imaging sensor, two transmission images are each simultaneously acquired by two linear CCD modules with different focal planes. Then the phase gradient information of the target can be obtained by an appropriate retrieval algorithm. Further, the multi-view three-dimensional effect is presented through dynamic angles of view. Theoretical analysis of this method is discussed, and experiments are carried out by building a scanner. The experiment results are presented with an algae specimen and transparent beads. We hope this method can be applied to present the three-dimensional effect of objects of flat translucent multilayer structure with a wide field of view.

Imaging moving targets through scattering media.

Optical microscopy in complex, inhomogeneous media is challenging due to the presence of multiply scattered light that limits the depths at which diffraction-limited resolution can be achieved. One way to circumvent the degradation in resolution is to use speckle- correlation-based imaging (SCI) techniques, which permit imaging of objects inside scattering media at diffraction-limited resolution. However, SCI methods are currently limited to imaging sparsely tagged objects in a dark-field scenario. In this work, we demonstrate the ability to image hidden, moving objects in a bright-field scenario. By using a deterministic phase modulator to generate a spatially incoherent light source, the background contribution can be kept constant between acquisitions and subtracted out. In this way, the signal arising from the object can be isolated, and the object can be reconstructed with high fidelity. With the ability to effectively isolate the object signal, our work is not limited to imaging bright objects in the dark-field case, but also works in bright-field scenarios, with non-emitting objects.

Focusing light through scattering media by transmission matrix inversion.

Focusing light through scattering media has broad applications in optical imaging, manipulation and therapy. The contrast of the focus can be quantified by peak-to-background intensity ratio (PBR). Here, we theoretically and numerically show that by using a transmission matrix inversion method to achieve focusing, within a limited field of view and under a low noise condition in transmission matrix measurements, the PBR of the focus can be higher than that achieved by conventional methods such as optical phase conjugation or feedback-based wavefront shaping. Experimentally, using a phase-modulation spatial light modulator, we increase the PBR by 66% over that achieved by conventional methods based on phase conjugation. In addition, we demonstrate that, within a limited field of view and under a low noise condition in transmission matrix measurements, our matrix inversion method enables light focusing to multiple foci with greater fidelity than those of conventional methods.

Quantitative phase imaging and complex field reconstruction by pupil modulation differential phase contrast.

Differential phase contrast (DPC) is a non-interferometric quantitative phase imaging method achieved by using an asymmetric imaging procedure. We report a pupil modulation differential phase contrast (PMDPC) imaging method by filtering a sample's Fourier domain with half-circle pupils. A phase gradient image is captured with each half-circle pupil, and a quantitative high resolution phase image is obtained after a deconvolution process with a minimum of two phase gradient images. Here, we introduce PMDPC quantitative phase image reconstruction algorithm and realize it experimentally in a 4f system with an SLM placed at the pupil plane. In our current experimental setup with the numerical aperture of 0.36, we obtain a quantitative phase image with a resolution of 1.73µm after computationally removing system aberrations and refocusing. We also extend the depth of field digitally by 20 times to ±50µm with a resolution of 1.76µm.

Analyzing the relationship between decorrelation time and tissue thickness in acute rat brain slices using multispeckle diffusing wave spectroscopy.

Novel techniques in the field of wavefront shaping have enabled light to be focused deep inside or through scattering media such as biological tissue. However, most of these demonstrations have been limited to thin, static samples since these techniques are very sensitive to changes in the arrangement of the scatterers within. As the samples of interest get thicker, the influence of the dynamic nature of the sample becomes even more pronounced and the window of time in which the wavefront solutions remain valid shrinks further. In this paper, we examine the time scales upon which this decorrelation happens in acute rat brain slices via multispeckle diffusing wave spectroscopy and investigate the relationship between this decorrelation time and the thickness of the sample using diffusing wave spectroscopy theory and Monte Carlo photon transport simulation.

Embedded pupil function recovery for Fourier ptychographic microscopy: erratum.

In the original paper, the line width of the resolution target (which corresponds to half-pitch resolution) was used to characterize the resolution of our microscope system. However, we think that full-pitch resolution offers a better definition of the imaging system's resolution limit. In this erratum, we list specific sections from the manuscript that used half-pitch resolution and correct them accordingly.

High numerical aperture Fourier ptychography: principle, implementation and characterization.

Fourier ptychography (FP) utilizes illumination control and computational post-processing to increase the resolution of bright-field microscopes. In effect, FP extends the fixed numerical aperture (NA) of an objective lens to form a larger synthetic system NA. Here, we build an FP microscope (FPM) using a 40X 0.75NA objective lens to synthesize a system NA of 1.45. This system achieved a two-slit resolution of 335 nm at a wavelength of 632 nm. This resolution closely adheres to theoretical prediction and is comparable to the measured resolution (315 nm) associated with a standard, commercially available 1.25 NA oil immersion microscope. Our work indicates that Fourier ptychography is an attractive method to improve the resolution-versus-NA performance, increase the working distance, and enlarge the field-of-view of high-resolution bright-field microscopes by employing lower NA objectives.

Solving ptychography with a convex relaxation.

Ptychography is a powerful computational imaging technique that transforms a collection of low-resolution images into a high-resolution sample reconstruction. Unfortunately, algorithms that currently solve this reconstruction problem lack stability, robustness, and theoretical guarantees. Recently, convex optimization algorithms have improved the accuracy and reliability of several related reconstruction efforts. This paper proposes a convex formulation of the ptychography problem. This formulation has no local minima, it can be solved using a wide range of algorithms, it can incorporate appropriate noise models, and it can include multiple a priori constraints. The paper considers a specific algorithm, based on low-rank factorization, whose runtime and memory usage are near-linear in the size of the output image. Experiments demonstrate that this approach offers a 25% lower background variance on average than alternating projections, the ptychographic reconstruction algorithm that is currently in widespread use.

A phase space model of Fourier ptychographic microscopy.

A new computational imaging technique, termed Fourier ptychographic microscopy (FPM), uses a sequence of low-resolution images captured under varied illumination to iteratively converge upon a high-resolution complex sample estimate. Here, we propose a mathematical model of FPM that explicitly connects its operation to conventional ptychography, a common procedure applied to electron and X-ray diffractive imaging. Our mathematical framework demonstrates that under ideal illumination conditions, conventional ptychography and FPM both produce datasets that are mathematically linked by a linear transformation. We hope this finding encourages the future cross-pollination of ideas between two otherwise unconnected experimental imaging procedures. In addition, the coherence state of the illumination source used by each imaging platform is critical to successful operation, yet currently not well understood. We apply our mathematical framework to demonstrate that partial coherence uniquely alters both conventional ptychography's and FPM's captured data, but up to a certain threshold can still lead to accurate resolution-enhanced imaging through appropriate computational post-processing. We verify this theoretical finding through simulation and experiment.

Axial standing-wave illumination frequency-domain imaging (SWIF).

Despite their tremendous contribution to biomedical research and diagnosis, conventional spatial sampling techniques such as wide-field, point scanning or selective plane illumination microscopy face inherent limiting trade-offs between spatial resolution, field-of-view, phototoxicity and recording speed. Several of these trade-offs are the result of spatial sampling with diffracting beams. Here, we introduce a new strategy for fluorescence imaging, SWIF, which instead encodes the axial profile of a sample in the Fourier domain. We demonstrate how this can be achieved with propagation-invariant illumination patterns that extend over several millimeters and robustly propagate through layers of varying refractive index. This enabled us to image a lateral field-of-view of 0.8 mm x 1.5 mm with an axial resolution of 2.4 µm - greatly exceeding the lateral field-of-view of conventional illumination techniques (~100 µm) at comparable resolution. Thus, SWIF allowed us to surpass the limitations of diffracting illumination beams and untangle lateral field-of-view from resolution.

Embedded pupil function recovery for Fourier ptychographic microscopy.

We develop and test a pupil function determination algorithm, termed embedded pupil function recovery (EPRY), which can be incorporated into the Fourier ptychographic microscopy (FPM) algorithm and recover both the Fourier spectrum of sample and the pupil function of imaging system simultaneously. This EPRY-FPM algorithm eliminates the requirement of the previous FPM algorithm for a priori knowledge of the aberration in the imaging system to reconstruct a high quality image. We experimentally demonstrate the effectiveness of this algorithm by reconstructing high resolution, large field-of-view images of biological samples. We also illustrate that the pupil function we retrieve can be used to study the spatially varying aberration of a large field-of-view imaging system. We believe that this algorithm adds more flexibility to FPM and can be a powerful tool for the characterization of an imaging system's aberration.

Model for estimating the penetration depth limit of the time-reversed ultrasonically encoded optical focusing technique.

The time-reversed ultrasonically encoded (TRUE) optical focusing technique is a method that is capable of focusing light deep within a scattering medium. This theoretical study aims to explore the depth limits of the TRUE technique for biological tissues in the context of two primary constraints - the safety limit of the incident light fluence and a limited TRUE's recording time (assumed to be 1 ms), as dynamic scatterer movements in a living sample can break the time-reversal scattering symmetry. Our numerical simulation indicates that TRUE has the potential to render an optical focus with a peak-to-background ratio of ~2 at a depth of ~103 mm at wavelength of 800 nm in a phantom with tissue scattering characteristics. This study sheds light on the allocation of photon budget in each step of the TRUE technique, the impact of low signal on the phase measurement error, and the eventual impact of the phase measurement error on the strength of the TRUE optical focus.