Sub-voxel light-sheet microscopy for high-resolution, high-throughput volumetric imaging of large biomedical specimens

Abstract

A key challenge when imaging whole biomedical specimens is how to quickly obtain massive cellular information over a large field of view (FOV). Here, we report a sub-voxel light-sheet microscopy (SLSM) method enabling high-throughput volumetric imaging of mesoscale specimens at cellular-resolution. A non-axial, continuous scanning strategy is used to rapidly acquire a stack of large-FOV images with three-dimensional (3-D) nanoscale shifts encoded. Then by adopting a sub-voxel-resolving procedure, the SLSM method models these low-resolution, cross-correlated images in the spatial domain and iteratively recovers a 3-D image with improved resolution throughout the sample. This technique can surpass the optical limit of a conventional light-sheet microscope by more than three times, with high acquisition speeds of gigavoxels per minute. As demonstrated by quick reconstruction (minutes to hours) of various samples, e.g., 3-D cultured cells, an intact mouse heart, mouse brain, and live zebrafish embryo, the SLSM method presents a high-throughput way to circumvent the tradeoff between intoto mapping of large-scale tissue (>100 mm3) and isotropic imaging of single-cell (~1-μm resolution). It also eliminates the need of complicated mechanical stitching or precisely modulated illumination, using a simple light-sheet setup and fast graphics-processing-unit (GPU)-based computation to achieve high-throughput, high-resolution 3-D microscopy, which could be tailored for a wide range of biomedical applications in pathology, histology, neuroscience, etc.

Simulating Developmental Cardiac Morphology in Virtual Reality Using a Deformable Image Registration Approach

“…Unlike our previous reports 11,12,15,32 , we have developed a resolution-enhancement method 16 with the retrospective synchronization algorithm for 4-D volumetric imaging of zebrafish embryos in a large field-of-view. Our novel pipeline was demonstrated linking deformable 3-D model architecture with the authentic dynamic LSFM data, and, founding the basis for recapitulating into a 4-D VR simulation of a contracting embryonic zebrafish heart.…”

Advances in Ex Situ Tissue Optical Clearing

“…Notably, both setups (Gaussian and Bessel) were characterized by similar lateral and axial full width at half maxima (1.56, 7.83 vs 1.77 and 8.47, respectively); however, utilization of Bessel beam was superior in all tested conditions, as it generated less striping artifacts (usually caused at the brain surface by bubbles or light absorption by bright structures), more homogenous image of deep brain structures, and a more complete picture of vasculature ( Figure ). In addition to optics, recent studies examined distinct arrangements of illumination and detection pathways in LSFM setup . Migliori et al .…”

Multiscale light-sheet for rapid imaging of cardiopulmonary system

“…Parallel advances in pixel superresolution (118)(119)(120), deep learning (121)(122)(123)(124), virtual reality (125)(126)(127)(128), optogenetics (129), and laser ablation (130) further provide complementary opportunities to elucidate cardiovascular architecture and function. A subvoxel LSFM is technically implemented for high-resolution, high-throughput volumetric imaging of cardiovascular development in a large field of view (22). This iterative resolution recovery method is transformative to improve inadequate focusing capability.…”

“…While PET (5,6), μCT (7,8), MRI (9,10), and bioluminescence imaging (11,12) are capable of capturing 3-D images from live samples, the spatial resolution of these techniques is inadequate to capture organ morphogenesis in small-animal models (13)(14)(15)(16)(17). For these reasons, the advent of lightsheet fluorescence microscopy (LSFM) (18)(19)(20)(21)(22) has revolutionized multiscale imaging, allowing visualization of samples ranging from live zebrafish embryos (~ 0.4 × 0.5 × 0.6 mm 3 ) to adult mouse hearts (~8 × 8 × 10 mm 3 ) with high-spatiotemporal resolution and minimal photobleaching and phototoxicity.…”