Super-resolved virtual staining of label-free tissue using diffusion models

Virtual staining of tissue offers a powerful tool for transforming label-free microscopy images of unstained tissue into equivalents of histochemically stained samples. This study presents a diffusion model-based super-resolution virtual staining approach utilizing a Brownian bridge process to enhance both the spatial resolution and fidelity of label-free virtual tissue staining, addressing the limitations of traditional deep learning-based methods. Our approach integrates novel sampling techniques into a diffusion model-based image inference process to significantly reduce the variance in the generated virtually stained images, resulting in more stable and accurate outputs. Blindly applied to lower-resolution auto-fluorescence images of label-free human lung tissue samples, the diffusion-based super-resolution virtual staining model consistently outperformed conventional approaches in resolution, structural similarity and perceptual accuracy, successfully achieving a super-resolution factor of 4-5x, increasing the output space-bandwidth product by 16-25-fold compared to the input label-free microscopy images. Diffusion-based super-resolved virtual tissue staining not only improves resolution and image quality but also enhances the reliability of virtual staining without traditional chemical staining, offering significant potential for clinical diagnostics.

Optical Generative Models

Generative models cover various application areas, including image, video and music synthesis, natural language processing, and molecular design, among many others. As digital generative models become larger, scalable inference in a fast and energy-efficient manner becomes a challenge. Here, we present optical generative models inspired by diffusion models, where a shallow and fast digital encoder first maps random noise into phase patterns that serve as optical generative seeds for a desired data distribution; a jointly-trained free-space-based reconfigurable decoder all-optically processes these generative seeds to create novel images (never seen before) following the target data distribution. Except for the illumination power and the random seed generation through a shallow encoder, these optical generative models do not consume computing power during the synthesis of novel images. We report the optical generation of monochrome and multi-color novel images of handwritten digits, fashion products, butterflies, and human faces, following the data distributions of MNIST, Fashion MNIST, Butterflies-100, and Celeb-A datasets, respectively, achieving an overall performance comparable to digital neural network-based generative models. To experimentally demonstrate optical generative models, we used visible light to generate, in a snapshot, novel images of handwritten digits and fashion products. These optical generative models might pave the way for energy-efficient, scalable and rapid inference tasks, further exploiting the potentials of optics and photonics for artificial intelligence-generated content.

BlurryScope: a cost-effective and compact scanning microscope for automated HER2 scoring using deep learning on blurry image data

We developed a rapid scanning optical microscope, termed "BlurryScope", that leverages continuous image acquisition and deep learning to provide a cost-effective and compact solution for automated inspection and analysis of tissue sections. BlurryScope integrates specialized hardware with a neural network-based model to quickly process motion-blurred histological images and perform automated pathology classification. This device offers comparable speed to commercial digital pathology scanners, but at a significantly lower price point and smaller size/weight, making it ideal for fast triaging in small clinics, as well as for resource-limited settings. To demonstrate the proof-of-concept of BlurryScope, we implemented automated classification of human epidermal growth factor receptor 2 (HER2) scores on immunohistochemically (IHC) stained breast tissue sections, achieving concordant results with those obtained from a high-end digital scanning microscope. We evaluated this approach by scanning HER2-stained tissue microarrays (TMAs) at a continuous speed of 5 mm/s, which introduces bidirectional motion blur artifacts. These compromised images were then used to train our network models. Using a test set of 284 unique patient cores, we achieved blind testing accuracies of 79.3% and 89.7% for 4-class (0, 1+, 2+, 3+) and 2-class (0/1+ , 2+/3+) HER2 score classification, respectively. BlurryScope automates the entire workflow, from image scanning to stitching and cropping of regions of interest, as well as HER2 score classification. We believe BlurryScope has the potential to enhance the current pathology infrastructure in resource-scarce environments, save diagnostician time and bolster cancer identification and classification across various clinical environments.

Lying mirror

We introduce an all-optical system, termed the "lying mirror", to hide input information by transforming it into misleading, ordinary-looking patterns that effectively camouflage the underlying image data and deceive the observers. This misleading transformation is achieved through passive light-matter interactions of the incident light with an optimized structured diffractive surface, enabling the optical concealment of any form of secret input data without any digital computing. These lying mirror designs were shown to camouflage different types of input image data, exhibiting robustness against a range of adversarial manipulations, including random image noise as well as unknown, random rotations, shifts, and scaling of the object features. The feasibility of the lying mirror concept was also validated experimentally using a structured micro-mirror array along with multi-wavelength illumination at 480, 550 and 600 nm, covering the blue, green and red image channels. This framework showcases the power of structured diffractive surfaces for visual information processing and might find various applications in defense, security and entertainment.

Deep Learning-based Detection of Bacterial Swarm Motion Using a Single Image

Distinguishing between swarming and swimming, the two principal forms of bacterial movement, holds significant conceptual and clinical relevance. This is because bacteria that exhibit swarming capabilities often possess unique properties crucial to the pathogenesis of infectious diseases and may also have therapeutic potential. Here, we report a deep learning-based swarming classifier that rapidly and autonomously predicts swarming probability using a single blurry image. Compared with traditional video-based, manually-processed approaches, our method is particularly suited for high-throughput environments and provides objective, quantitative assessments of swarming probability. The swarming classifier demonstrated in our work was trained on Enterobacter sp. SM3 and showed good performance when blindly tested on new swarming (positive) and swimming (negative) test images of SM3, achieving a sensitivity of 97.44% and a specificity of 100%. Furthermore, this classifier demonstrated robust external generalization capabilities when applied to unseen bacterial species, such as Serratia marcescens DB10 and Citrobacter koseri H6. It blindly achieved a sensitivity of 97.92% and a specificity of 96.77% for DB10, and a sensitivity of 100% and a specificity of 97.22% for H6. This competitive performance indicates the potential to adapt our approach for diagnostic applications through portable devices or even smartphones. This adaptation would facilitate rapid, objective, on-site screening for bacterial swarming motility, potentially enhancing the early detection and treatment assessment of various diseases, including inflammatory bowel diseases (IBD) and urinary tract infections (UTI).

Label-free evaluation of lung and heart transplant biopsies using virtual staining

Organ transplantation serves as the primary therapeutic strategy for end-stage organ failures. However, allograft rejection is a common complication of organ transplantation. Histological assessment is essential for the timely detection and diagnosis of transplant rejection and remains the gold standard. Nevertheless, the traditional histochemical staining process is time-consuming, costly, and labor-intensive. Here, we present a panel of virtual staining neural networks for lung and heart transplant biopsies, which digitally convert autofluorescence microscopic images of label-free tissue sections into their brightfield histologically stained counterparts, bypassing the traditional histochemical staining process. Specifically, we virtually generated Hematoxylin and Eosin (H&E), Masson's Trichrome (MT), and Elastic Verhoeff-Van Gieson (EVG) stains for label-free transplant lung tissue, along with H&E and MT stains for label-free transplant heart tissue. Subsequent blind evaluations conducted by three board-certified pathologists have confirmed that the virtual staining networks consistently produce high-quality histology images with high color uniformity, closely resembling their well-stained histochemical counterparts across various tissue features. The use of virtually stained images for the evaluation of transplant biopsies achieved comparable diagnostic outcomes to those obtained via traditional histochemical staining, with a concordance rate of 82.4% for lung samples and 91.7% for heart samples. Moreover, virtual staining models create multiple stains from the same autofluorescence input, eliminating structural mismatches observed between adjacent sections stained in the traditional workflow, while also saving tissue, expert time, and staining costs.

Unidirectional imaging with partially coherent light

Unidirectional imagers form images of input objects only in one direction, e.g., from field-of-view (FOV) A to FOV B, while blocking the image formation in the reverse direction, from FOV B to FOV A. Here, we report unidirectional imaging under spatially partially coherent light and demonstrate high-quality imaging only in the forward direction (A->B) with high power efficiency while distorting the image formation in the backward direction (B->A) along with low power efficiency. Our reciprocal design features a set of spatially engineered linear diffractive layers that are statistically optimized for partially coherent illumination with a given phase correlation length. Our analyses reveal that when illuminated by a partially coherent beam with a correlation length of ~1.5 w or larger, where w is the wavelength of light, diffractive unidirectional imagers achieve robust performance, exhibiting asymmetric imaging performance between the forward and backward directions - as desired. A partially coherent unidirectional imager designed with a smaller correlation length of less than 1.5 w still supports unidirectional image transmission, but with a reduced figure of merit. These partially coherent diffractive unidirectional imagers are compact (axially spanning less than 75 w), polarization-independent, and compatible with various types of illumination sources, making them well-suited for applications in asymmetric visual information processing and communication.

Virtual Gram staining of label-free bacteria using darkfield microscopy and deep learning

Gram staining has been one of the most frequently used staining protocols in microbiology for over a century, utilized across various fields, including diagnostics, food safety, and environmental monitoring. Its manual procedures make it vulnerable to staining errors and artifacts due to, e.g., operator inexperience and chemical variations. Here, we introduce virtual Gram staining of label-free bacteria using a trained deep neural network that digitally transforms darkfield images of unstained bacteria into their Gram-stained equivalents matching brightfield image contrast. After a one-time training effort, the virtual Gram staining model processes an axial stack of darkfield microscopy images of label-free bacteria (never seen before) to rapidly generate Gram staining, bypassing several chemical steps involved in the conventional staining process. We demonstrated the success of the virtual Gram staining workflow on label-free bacteria samples containing Escherichia coli and Listeria innocua by quantifying the staining accuracy of the virtual Gram staining model and comparing the chromatic and morphological features of the virtually stained bacteria against their chemically stained counterparts. This virtual bacteria staining framework effectively bypasses the traditional Gram staining protocol and its challenges, including stain standardization, operator errors, and sensitivity to chemical variations.

Multi-scale Conditional Generative Modeling for Microscopic Image Restoration

The advance of diffusion-based generative models in recent years has revolutionized state-of-the-art (SOTA) techniques in a wide variety of image analysis and synthesis tasks, whereas their adaptation on image restoration, particularly within computational microscopy remains theoretically and empirically underexplored. In this research, we introduce a multi-scale generative model that enhances conditional image restoration through a novel exploitation of the Brownian Bridge process within wavelet domain. By initiating the Brownian Bridge diffusion process specifically at the lowest-frequency subband and applying generative adversarial networks at subsequent multi-scale high-frequency subbands in the wavelet domain, our method provides significant acceleration during training and sampling while sustaining a high image generation quality and diversity on par with SOTA diffusion models. Experimental results on various computational microscopy and imaging tasks confirm our method's robust performance and its considerable reduction in its sampling steps and time. This pioneering technique offers an efficient image restoration framework that harmonizes efficiency with quality, signifying a major stride in incorporating cutting-edge generative models into computational microscopy workflows.

Integration of Programmable Diffraction with Digital Neural Networks

Optical imaging and sensing systems based on diffractive elements have seen massive advances over the last several decades. Earlier generations of diffractive optical processors were, in general, designed to deliver information to an independent system that was separately optimized, primarily driven by human vision or perception. With the recent advances in deep learning and digital neural networks, there have been efforts to establish diffractive processors that are jointly optimized with digital neural networks serving as their back-end. These jointly optimized hybrid (optical+digital) processors establish a new "diffractive language" between input electromagnetic waves that carry analog information and neural networks that process the digitized information at the back-end, providing the best of both worlds. Such hybrid designs can process spatially and temporally coherent, partially coherent, or incoherent input waves, providing universal coverage for any spatially varying set of point spread functions that can be optimized for a given task, executed in collaboration with digital neural networks. In this article, we highlight the utility of this exciting collaboration between engineered and programmed diffraction and digital neural networks for a diverse range of applications. We survey some of the major innovations enabled by the push-pull relationship between analog wave processing and digital neural networks, also covering the significant benefits that could be reaped through the synergy between these two complementary paradigms.