Recovering molecular information remains a grand challenge in the widely used holographic and computational imaging technologies. To address this challenge, we developed a computational mid-infrared photothermal microscope, termed Bond-selective Intensity Diffraction Tomography (BS-IDT). Based on a low-cost brightfield microscope with an add-on pulsed light source, BS-IDT recovers both infrared spectra and bond-selective 3D refractive index maps from intensity-only measurements. High-fidelity infrared fingerprint spectra extraction is validated. Volumetric chemical imaging of biological cells is demonstrated at a speed of ~20 s per volume, with a lateral and axial resolution of ~350 nm and ~1.1 µm, respectively. BS-IDT’s application potential is investigated by chemically quantifying lipids stored in cancer cells and volumetric chemical imaging on Caenorhabditis elegans with a large field of view (~100 µm x 100 µm).
We develop a novel algorithm for large-scale holographic reconstruction of 3D particle fields. Our method is based on a multiple-scattering beam propagation method (BPM) combined with sparse regularization that enables recovering dense 3D particles of high refractive index contrast from a single hologram. We show that the BPM-computed hologram generates intensity statistics closely matching with the experimental measurements and provides up to 9× higher accuracy than the single-scattering model. To solve the inverse problem, we devise a computationally efficient algorithm, which reduces the computation time by two orders of magnitude as compared to the state-of-the-art multiple-scattering based technique. We demonstrate the superior reconstruction accuracy in both simulations and experiments under different scattering strengths. We show that the BPM reconstruction significantly outperforms the single-scattering method in particular for deep imaging depths and high particle densities.
Segmentation of blood vessels from two-photon microscopy (2PM) angiograms of brains has important applications in hemodynamic analysis and disease diagnosis. Here we develop a generalizable deep-learning technique for accurate 2PM vascular segmentation of sizable regions in mouse brains acquired from multiple 2PM setups. In addition, the technique is computationally efficient, making it ideal for large-scale neurovascular analysis. Introduction: Vascular segmentation from 2PM angiograms is usually an important first step in hemodynamic modeling of brain vasculature. Existing state-of-the-art segmentation methods based on deep learning either lack the ability to generalize to data from various imaging systems, or are computationally infeasible for large-scale angiograms. In this work, we present a method which improves upon both these limitations by being generalizable to various imaging systems, and also being able to segment very large-scale angiograms. Methods: We employ a computationally efficient deep learning framework based on a semi-supervised learning strategy, whose effectiveness we demonstrate on experimentally acquired in-vivo angiograms from mouse brains of dimensions up to 808x808x702 micrometers. Results: After training on data from only one 2PM microscope, we perform vascular segmentation on data from another microscope without any network tuning. Our method demonstrates 10x faster computation in terms of voxels-segmented-persecond and 3x larger depth compared to the state-of-the-art. Conclusion: Our work provides a generalizable and computationally efficient anatomical modeling framework for the brain vasculature, which consists of deeplearning based vascular segmentation followed by graphing. It paves the way for future modeling and analysis of hemodynamic response at much greater scales that were inaccessible before.
Objective and Impact Statement. Segmentation of blood vessels from two-photon microscopy (2PM) angiograms of brains has important applications in hemodynamic analysis and disease diagnosis. Here, we develop a generalizable deep learning technique for accurate 2PM vascular segmentation of sizable regions in mouse brains acquired from multiple 2PM setups. The technique is computationally efficient, thus ideal for large-scale neurovascular analysis. Introduction. Vascular segmentation from 2PM angiograms is an important first step in hemodynamic modeling of brain vasculature. Existing segmentation methods based on deep learning either lack the ability to generalize to data from different imaging systems or are computationally infeasible for large-scale angiograms. In this work, we overcome both these limitations by a method that is generalizable to various imaging systems and is able to segment large-scale angiograms. Methods. We employ a computationally efficient deep learning framework with a loss function that incorporates a balanced binary-cross-entropy loss and total variation regularization on the network’s output. Its effectiveness is demonstrated on experimentally acquired in vivo angiograms from mouse brains of dimensions up to 808×808×702 μm. Results. To demonstrate the superior generalizability of our framework, we train on data from only one 2PM microscope and demonstrate high-quality segmentation on data from a different microscope without any network tuning. Overall, our method demonstrates 10× faster computation in terms of voxels-segmented-per-second and 3× larger depth compared to the state-of-the-art. Conclusion. Our work provides a generalizable and computationally efficient anatomical modeling framework for brain vasculature, which consists of deep learning-based vascular segmentation followed by graphing. It paves the way for future modeling and analysis of hemodynamic response at much greater scales that were inaccessible before.
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