Cores for Vision Research Administration Bioinformatics Core Ocular Microanatomy Core (OM Core) Visual Function and Non-invasive Ocular Imaging Core (VFNOI Core) Ocular Mass Spectrometry, Lipidomics, and Proteomics Core (OMSLP Core) Ophthalmic Testing Core (OTC) Ocular Microanatomy Core (OM Core) Home Research Cores For Vision Research Ocular Microanatomy Core (OM Core) Ocular Microanatomy Core (OM Core) Cores of Vision Research within the University of California Irvine (UCI) will strengthen and enhance NEI R01-funded research. The project aims to support the major expansion of vision research currently underway at UCI. Our goal is to broaden and optimize research capabilities, support and expand existing research projects, and help establish new studies, ultimately boosting productivity and strengthening vision research at UCI. It is sponsored by the NEI P30 grant, P30EY034070-01. For feedback please email Dr. Zode or Dr. Linya Li Ocular Microanatomy Core Specimen submission form Gulab Zode, PhD Core Director, Ocular Microanatomy Core Professor, Gavin Herbert Eye Institute Linya Li Core Manager, Ocular Microanatomy Core Core Overview The Ocular Microanatomy Core (OM Core) provides centralized histology, advanced fluorescence imaging, and quantitative image-analysis support for NEI-funded investigators and the broader UCI vision-research community. Supported by NEI Center Core Grant P30EY034070-01, the Core strengthens and expands the research infrastructure needed for the major growth of vision research underway at UCI. Our goal is to broaden and optimize research capabilities, accelerate existing projects, enable new studies, and improve the rigor, reproducibility, and productivity of ocular research. Core staff provide project consultation, standardized sample preparation, access to specialized instrumentation, hands-on training, assisted imaging, and support for quantitative analysis. Services available The OM Core supports projects from ocular-tissue preparation through image acquisition and analysis. Services can be requested individually or combined into an end-to-end workflow. Project consultation: Selection of fixation, embedding, section thickness, staining, imaging modality, and analysis endpoints. Frozen histology: OCT embedding, controlled freezing, cryostat sectioning, serial or step-section collection, slide preparation, and storage guidance. Paraffin histology: Automated tissue processing, paraffin embedding, rotary-microtome sectioning, water-bath transfer, and slide preparation. Routine staining: Hematoxylin and eosin (H&E). Fluorescence imaging: High-sensitivity confocal imaging, optical sectioning, z-stacks, tiled mosaics, multichannel acquisition, and three-dimensional visualization. Two-photon imaging: Multiphoton excitation for deeper imaging of thick specimens and intact or whole-mount ocular preparations. Live-cell imaging: Time-lapse imaging with environmental control for temperature and CO₂, supporting dynamic studies in cultured cells and viable specimens. Image analysis: Leica software-supported stitching, z-stack review, 3D rendering, intensity measurements, colocalization, cell counting, morphometry, and figure-ready export, as appropriate to the project. Typical Project Workflow Discuss the biological question, specimen type, orientation, experimental groups, and quantitative endpoints with the Core Manager. Confirm the fixation, OCT or paraffin workflow, sectioning strategy, stains or fluorescent labels, and imaging requirements. Submit clearly labeled samples with a sample manifest and requested sectioning or imaging plan. Review pilot sections or images before full-batch processing when optimization is needed. Receive slides and/or image data with guidance for data organization, analysis, and interpretation. Major Equipment Leica CM1860 Cryostat Used to section fresh or fixed ocular tissues embedded in OCT medium. The cryostat produces reproducible serial or step sections for routine histology, immunofluorescence, and other downstream applications. Leica CM1860 cryostat for OCT-embedded ocular tissues. Nikon SMZ745 Stereomicroscope Supports ocular tissue dissection, specimen orientation, and preparation of whole eyes, retina, optic nerve, anterior segment, and other small tissue samples. Nikon SMZ745 stereomicroscope for ocular dissection and orientation Paraffin Histology Leica TP1020 Automated Tissue Processor Provides automated dehydration, clearing, and paraffin infiltration of fixed ocular tissues for consistent preparation of paraffin blocks. Leica TP1020 automated tissue processor. Paraffin Embedding Workstation A heated embedding center with a cold module enables controlled specimen orientation and uniform paraffin-block preparation before sectioning. Paraffin embedding workstation with heated and cold modules. Leica TP1020 Automated Tissue Processor Provides automated dehydration, clearing, and paraffin infiltration of fixed ocular tissues for consistent preparation of paraffin blocks. Leica rotary microtome and water-bath system for paraffin sectioning. Dedicated Histology Staining Station Supports standardized H&E staining of ocular sections. PAS, Masson’s trichrome, and other conventional stains are available by consultation. Dedicated H&E staining station in the FLF 6525 histology laboratory. Representative output: H&E-stained ocular section. Leica STELLARIS Confocal and Multiphoton Imaging System Provides high-sensitivity, multichannel fluorescence imaging of cultured cells, tissue sections, thick specimens, and retinal whole mounts. Capabilities include confocal optical sectioning, z-stacks, tiled imaging, image stitching, 3D visualization, and two-photon imaging of deeper tissue structures. Leica STELLARIS confocal microscope system with enclosed imaging chamber. Live-Cell Environmental Control Temperature- and CO₂-controlled incubation supports time-lapse and live-cell fluorescence imaging under physiologically controlled conditions. Merged multichannel retinal whole-mount image showing broad tissue coverage and cellular detail. Representative single-channel view from the same retinal whole-mount dataset. Leica Image-Acquisition and Analysis Workstation Provides microscope control, multichannel acquisition, tile stitching, z-stack processing, 3D visualization, quantitative measurements, and image export using Leica software. Dedicated Leica acquisition and image-analysis workstation. Module services and equipment available Routine Paraffin Embedding, Sectioning and Staining. At present, this is a partial service where investigators submit tissue to outside parties for paraffin embedding. Paraffin blocks are then returned for sectioning and staining within the Core. Paraffin blocks will be cut using a Leica RM 2125 Rotary Microtome and section placed on glass slides. The number of slides needed per sample, including serial or step sectioning through specific structures should be communicated to the OMC Manager. Standard staining of tissue sections can also be requested, including Hematoxylin & Eosin, Periodic Acid Schiff, and Masson’s Trichrome. Investigators interested in performing immunohistochemistry will receive unstained slides for performing immunostaining in their own laboratories. We also have available a TedPella Biowave for antigen retrieval and rapid immunostaining of paraffin processed tissue. Immunofluorescence This service provides embedding of fixed or fresh submitted tissue for Optimal Cutting Temperature media embedding, snap freezing in liquid nitrogen/isopentane and tissue storage at -80oC. Tissue will then be sectioned on a Leica CM 1850 Cryostat, and unstained tissue sections provided to the investigator for immunostaining in their own laboratories. We also have the capability of performing immunofluorescent staining generating thick tissue sections either with a Campden Instruments Ci 5100 mz Vibratome. Immuno-Tomography The OMC offers also Immuno-tomography (IT), a new and innovative technology that allows 3-D reconstruction of tissues. IT is a powerful 3-dimensional reconstruction technique where large tissue blocks (2 – 3 mm across) are embedded in Butyl Methyl Methacrylate (BMMA) and then serial sectioned at 2 μm thickness using an Ultramicrotome. Up to 500 sections (1 mm into the block) are floated onto standard glass slides for immunostaining. As shown below, tissue is embedded in plastic, then serial sectioned using a Leica EM UC7 Ultramicrotome, and \tissue sections imaged using a tiling routine on our automated inverted fluorescent microscope (Leica DMI6000B Fluorescent Inverted Microscope. Images are automatically stitched and then the tissue 3 dimensionally reconstructed at submicron resolution using Amira software. Slides are then stripped of antibodies and the process is repeated with different probes to yield a 3D representation of the tissue or cells of interest, with emphasis on immunochemically identified structures (illustrated below). Image Processing Images generated from any of the microscope devices are analyzed on one of two PCs that have dual Intel Xeon Quad Core CPUs clocked at 2.5 GHz per core with 16 GB of RAM and a NVIDIA GeForce GTK-46 graphic card with 9G of memory. Both computers are networked to our two 5 Terabyte QNAP Turbo NAS online back-up drives for safely storing images while processing. Both computers also run the same image processing software including AMIRA 5.4.3 (Visage Imaging, Carlsbad, CA) and Metamorph Offline Version 7.8.6.0 (Molecular Devices, San Jose, CA) as well as NIH ImageJ and other free image processing software. Training on the use of these programs for image processing, quantitative image analysis and 3 dimensional reconstruction will be provided by the Microscopy and Imaging Manager.