Isolation of Intact Eyeball to Obtain Integral Ocular Surface Tissue for Histological Examination and Immunohistochemistry
Isolation of Intact Eyeball to Obtain Integral Ocular Surface Tissue for Histological Examination and Immunohistochemistry
复制标题
分离完整的眼球以获得完整的眼表组织用于组织学检查和免疫组织化学
DOI：
10.3791/60086
复制
发表时间：
2019-10-01
影响因子：
1.2
通讯作者：
Liu, Chunqiao
中科院分区：
文献类型：
--
作者：
Guo, Dianlei;Liu, Chunqiao
眼表（OS）由三个相连部分组成：眼睑结膜、球结膜和角膜上皮。破坏OS会导致角膜炎、结膜炎或两者兼而有之（角膜结膜炎）。在经过一定遗传修饰或人工操作的实验动物模型中，检查OS上皮片的所有部分，并行评估各部分的相对发病变化是有用的。然而，在不变形或不损伤的情况下解剖整个OS组织一直是一项挑战，这主要是由于附着在物理上分离但可活动的眼睑和眼球上的OS柔软而薄。此外，由硬颅骨/眶骨形成的深眼窝被眼球完全占据，留给手术解剖的空间有限。因此，直接从面侧剥离眼球及相关的OS组织往往会导致组织损伤，尤其是眼睑和球结膜。在本方案中，我们描述了一种从一分为二的小鼠头部依次去除颅骨和眶骨的方法，留下眼睑，眼表，晶状体和视网膜在一起。OS片的完整性得到很好的保存，可以通过组织学或免疫染色在单个切片中进行检查。
Ocular surface (OS) consists of an epithelial sheet with three connected parts: palpebral conjunctiva, bulbar conjunctiva and corneal epithelium. Disruption of OS would lead to keratitis, conjunctivitis or both (keratoconjunctivitis). In experimental animal models with certain genetic modifications or artificial operations, it is useful to examine all parts of the OS epithelial sheet to evaluate relative pathogenetic changes of each part in parallel. However, dissection of OS tissue as a whole without distortion or damage has been challenging, primarily due to the softness and thinness of the OS affixed to physically separate yet movable eyelids and eyeball. Additionally, the deep eye socket formed by the hard skull/orbital bones is fully occupied by the eyeball leaving limited space for operating dissections. As a result, direct dissection of the eyeball with associated OS tissues from the facial side would often lead to tissue damages, especially the palpebral and bulbar conjunctiva. In this protocol, we described a method to remove the skull and orbital bones sequentially from a bisected mouse head, leaving eyelids, ocular surface, lens and retina altogether in one piece. The integrity of the OS sheet was well preserved and could be examined by histology or immunostaining in a single section.