Histocore arcadia embedding center

The livers of controls and TAA-treated rats were removed and fixed in 10% formalin, regularly subjected to paraffin sections, and stained with hematoxylin and eosin (H and E). Briefly, after gradient dehydration with various concentrations of alcohol in an automatic tissue dehydrator (HistoCore PELORIS 3 Premium Tissue Processing System, Leica Biosystems Inc., Buffalo Grove, IL, USA), the liver of each animal was embedded in paraffin blocks (using HistoCore Arcadia Embedding Center, Leica Biosystems Inc., Buffalo Grove, IL, USA). The liver tissue was then cut into 10 µm thin slices via an ultra-thin semiautomatic microtome (Histocoreautocut automated rotary microtome, Leica Biosystems Inc., Buffalo Grove, IL, USA), and adhered to the slides. Then, the slides were stained with H and E, and the morphological changes were evaluated using a microscope (Olympus VS120 Automated Slide Scanner, Olympus, Pittsburgh, PA, USA) [25 (link),26 (link)].

Eyes were enucleated and fixed in 4% paraformaldehyde (AR1068; Boster Bio, Pleasanton, CA, USA) for 48 hours at 4°C. Whole eyeballs were dehydrated and infiltrated with paraffin using a tissue processor (Tissue-Tek VIP 6; Sakura Finetek USA, Inc., Torrance, CA, USA) and subsequently embedded into paraffin blocks on a HistoCore Arcadia Embedding Center (Leica Biosystems, Melbourne, Australia). Three-micrometer sections were cut at the level of the optical nerve and hematoxylin and eosin and Masson's Trichrome staining was performed according to standard protocols. Immunofluorescence staining was carried out on the automated Leica Bond platform (Leica Biosystems) using the Opal Multiplex IHC Kit (Akoya Biosciences, Menlo Park, CA, USA). Antibodies used in this study and details about the antigen retrieval are listed in the Table. Nuclei were stained with spectral DAPI (FP1490; Akoya Biosciences) and slides mounted with ProLong Antifade Mounting Medium (P36961; Invitrogen, Carlsbad, CA, USA). Tissues were imaged using an Axio Scan.Z1 slide scanner (magnification ×20; Carl Zeiss Microscopy GmbH, Jena, Germany). Iba1⁺ and F4/80⁺ area was quantified using HALO image analysis software (Indica Labs, Albuquerque, NM, USA).