Optimized technique to extract and fix Olive ridley turtle hatchling retina for histological study

Abstract The efficient extraction and fixation of a tissue allows in preserving the cytoarchitecture, chemical composition and tissue organization, which is key in physiological and histopathological studies. The main goal of this study was to establish a microsurgery technique to obtain ocular tissue and provide an optimized immersion fixation protocol based on the 10 % formalin-intraocular injection on Olive ridley sea turtle hatchlings (Lepidochelys olivacea). To evaluate this optimized technique, a histological comparison between traditional immersion and intraocular/immersion protocols was done. The eyeball were processed into five protocols: Frozen eyes (Group 1), frozen eyes immersed in 10 % formalin (Group 2), fresh eyes immersed in 10 % formalin (Group 3), fresh eyes intraocularly injected with 0.1 M phosphate buffer solution (PBS) and then immersed in 10 % formalin (Group 4), and fresh eyes fixed by 10 % formalin-intraocular followed by 10 % formalin-immersion (Group 5). In comparison with all groups evaluated, the intraocular/immersion fixation protocol lead the conservation of eyeball shape, cell integrity and maintenance of the organization of the retina layers of sea turtle hatchlings. If this method will be the key in studying sea turtle, we suppose that this procedure, with minimal adjustments, could be useful in animals with similar eye anatomy.