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使用新型单克隆抗体RT10F7对大鼠视网膜中穆勒细胞的发育研究

Developmental study of Müller cells in the rat retina using a new monoclonal antibody, RT10F7.

作者信息

Yamasaki E N, Krupnik V E, Chun L L

机构信息

Retinal Development and Regeneration Laboratory, Neurosurgical Service, Massachusetts General Hospital, Harvard Medical School, Boston, USA.

出版信息

Neuroscience. 1998 Jul;85(2):627-36. doi: 10.1016/s0306-4522(97)00084-5.

Abstract

We produced the monoclonal antibody RT10F7, characterized its antigenic specificity and expression in the adult and developing retina, in cultured retinal cells and in other parts of the central nervous system. In metabolically-labelled retinal cultures RT10F7 immunoprecipitated a protein of approximately 36,000 mol. wt. In the adult, RT10F7 stained endfeet of Müller cells in the ganglion cell layer, four horizontal bands in the inner plexiform layer, and radial fibres in the outer plexiform layer which terminated at the outer limiting membrane. In the inner nuclear layer, most somata were underlined by Müller processes that wrapped around them, but some cell bodies were immunoreactive for RT10F7 in the cytoplasm. During development, postnatal day 21 was the first age at which the adult pattern of immunoreactivity was present, although a fourth band in the inner plexiform layer was less clear than for the adult. By 14 and eight days after birth, the pattern of RT10F7 immunoreactivity approximated that of the adult; however, only three bands and one band were present, respectively, in the inner plexiform layer. At earlier ages, postnatal days 4, 1 and embryonic ages 19 and 15, the monoclonal antibody stained Müller cell endfeet and radial fibres, from the inner plexiform layer through the neuroblastic layer to the outer limiting membrane. At these ages, the immunoreactivity was more prominent at the level of Müller cell endfeet. The monoclonal antibody stained glia in preparations of dissociated retinal cells maintained in culture but not astrocytes or oligodendrocytes from optic nerve cultures. In brain sections, tanycytes exhibited RT10F7 immunoreactivity. The monoclonal antibody RT10F7 recognized a specific cell type in the retina, the Müller cell. In the adult and developing retina, RT10F7 recognized an antigen that is present primarily in Müller cell processes. This feature allowed us to follow the maturation of the Müller cell and correlate it with developmental events in the retina. RT10F7 is a specific marker for Müller cells in vivo and in vitro and may be useful for studies of function of Müller cells after ablation or after injuries that are known to activate Müller cells.

摘要

我们制备了单克隆抗体RT10F7,对其抗原特异性进行了表征,并研究了其在成年和发育中的视网膜、培养的视网膜细胞以及中枢神经系统其他部位的表达情况。在代谢标记的视网膜培养物中,RT10F7免疫沉淀出一种分子量约为36,000的蛋白质。在成体中,RT10F7染色显示,神经节细胞层中Müller细胞的终足、内网状层中的四条水平带以及外网状层中终止于外限制膜的放射状纤维均被染色。在内核层中,大多数细胞体被环绕它们的Müller细胞突起所覆盖,但有些细胞体的细胞质对RT10F7呈免疫反应性。在发育过程中,出生后第21天是首次出现成年免疫反应模式的年龄,尽管内网状层中的第四条带不如成体清晰。出生后14天和8天时,RT10F7免疫反应模式接近成年模式;然而,内网状层中分别仅出现三条带和一条带。在更早的年龄,出生后第4天、第1天以及胚胎期第19天和第15天,单克隆抗体染色显示,Müller细胞终足和放射状纤维,从内网状层穿过神经母细胞层直至外限制膜均被染色。在这些年龄,免疫反应性在Müller细胞终足水平更为突出。单克隆抗体在培养的解离视网膜细胞制剂中对神经胶质细胞染色,但对视神经培养物中的星形胶质细胞或少突胶质细胞不染色。在脑切片中,伸展细胞呈现RT10F7免疫反应性。单克隆抗体RT10F7识别视网膜中的一种特定细胞类型,即Müller细胞。在成年和发育中的视网膜中,RT10F7识别一种主要存在于Müller细胞突起中的抗原。这一特性使我们能够追踪Müller细胞的成熟过程,并将其与视网膜中的发育事件相关联。RT10F7是体内和体外Müller细胞的特异性标志物,可能有助于研究Müller细胞在被切除或在已知会激活Müller细胞的损伤后的功能。

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