Abstract
Purpose :
Conventional histology faces limitations due to sectioning artifacts and three-dimensional misrepresentation, impeding direct comparison with clinical imaging. The ClearEye method involves clearing entire human eyes for examination through light sheet fluorescence microscopy (LSFM), enabling comprehensive 3D imaging of complete donor eyes while preserving spatial architecture. ClearEye allows customizable observations, including en face and transverse scans, showcasing diverse capabilities in 3D navigation and virtual dissection.
Methods :
Seventeen ocular samples from healthy donors (age 16 to 96 years) underwent immunolabeling (tubulin III, collagen IV, alpha-SMA antibodies) and clearing with the ClearEye protocol. Cleared samples were imaged using a custom-designed LSFM (MesoSPIM), and 3D analysis was performed with Imaris software.
Results :
ClearEye-cleared tissues facilitated exploration of spatially complex structures through virtual navigation. En face and transverse viewing replicated orientations similar to slit lamp, fundus photographs, or OCT scans, aiding clinicians in data interpretation. Virtual dissection isolated specific structures such as the retinal nerve fiber layer (RNFL), choroid, Schlemm's canal, and anterior ciliary arteries. The intraneural path of the central retinal artery and vein, and the Zinn-Haller circle, became observable. Capillary connections in the optic nerve revealed dual arterial perfusion (pial and central artery), with predominant venous drainage via the central retinal vein. Analysis of age-related changes in the anterior segment demonstrated impacts on nerve fibers in the cornea, iris, and ciliary bodies.
Conclusions :
In conclusion, 3D histology presents a groundbreaking, multi-scale approach, enhancing understanding of ocular diseases. It improves comprehension of perfusion schemes in structures traditionally inaccessible to imaging, establishing a synergistic bridge between clinical imaging and histological data. This integrated approach reveals crucial structural-functional relationships, laying the foundation for precise characterization of pathologies, particularly beneficial for myopia, vitreous diseases, and plurifocal diseases like uveitis.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.