Ariyasu R G, Nichol J A, Ellisman M H
J Neurosci. 1985 Oct;5(10):2581-96. doi: 10.1523/JNEUROSCI.05-10-02581.1985.
This report describes the development and characterization of a battery of highly specific antibodies to sodium/potassium (Na+ + K+)-ATPase and their use in localizing this enzyme in nervous tissue. The immunolabeling characteristics of polyclonal antibodies and monoclonal antibodies (Schenk, D. B., and H. L. Leffert (1983) Proc. Natl. Acad. Sci. U. S. A. 80: 5281-5285) raised against rat renal (Na+ + K+)-ATPase were compared. The interspecies cross-reactivity of the polyclonal anti-rat antibodies was examined by determining their binding to purified rat, eel, or dog enzyme. The immunostaining characteristics of the IgG fraction of the polyclonal antibody preparations, their affinity-purified derivatives, and the monoclonal antibodies were compared. The results obtained with each of these were similar, providing information about where focal concentrations of the enzyme exist within nervous tissue. The IgG fraction of the polyclonal antibody preparations provided the most sensitive probe, facilitating localization of the (Na+ + K+)-ATPase in the tissue sections from various regions of the nervous system. (Na+ + K+)-ATPase-like immunoreactivity was observed along the plasmalemma of alpha-motor neurons and at the nodal axolemma of myelinated axons from the central or peripheral nervous system. It was determined that the absence of labeling for the enzyme along the paranodal or internodal regions of the axolemma was not an artifact due to a limited accessibility of antibody to these regions. Some central nervous system glial cells demonstrated abundant amounts of plasmalemmal and intracellular (Na+ + K+)-ATPase-like immunoreactivity. These cells were identified as astroglia by positive labeling of cells in serial sections for glial fibrillary acid protein immunoreactivity in the soma and radial processes in optic nerve, or velous processes in the cerebellum. Astrocyte processes overlying the nodal axolemma also stained positively for the enzyme. (Na+ + K+)-ATPase-like immunoreactivity was not observed in association with oligodendroglia cell bodies or their processes forming myelin sheaths. In contrast, the plasmalemma of myelinating Schwann cells showed greatest immunoreactivity in the region of the node of Ranvier. Although a focal concentration of immunoreactivity was observed along node- and paranode-associated regions of Schwann cells, a lower level of uniform staining was noted along the entire Schwann cell surface membrane.(ABSTRACT TRUNCATED AT 400 WORDS)
本报告描述了一系列针对钠/钾（Na⁺+K⁺）-ATP酶的高特异性抗体的研制及其特性，并阐述了这些抗体在神经组织中定位该酶的应用。比较了针对大鼠肾（Na⁺+K⁺）-ATP酶产生的多克隆抗体和单克隆抗体（申克，D.B.，和H.L.莱弗特（1983年）《美国国家科学院院刊》80:5281 - 5285）的免疫标记特性。通过测定多克隆抗大鼠抗体与纯化的大鼠、鳗鱼或犬类酶的结合情况，研究了其种间交叉反应性。比较了多克隆抗体制剂的IgG组分、亲和纯化衍生物以及单克隆抗体的免疫染色特性。这些抗体各自获得的结果相似，提供了有关该酶在神经组织中局部浓度存在位置的信息。多克隆抗体制剂的IgG组分提供了最敏感的探针，便于在来自神经系统各个区域的组织切片中定位（Na⁺+K⁺）-ATP酶。在α运动神经元的质膜以及中枢或外周神经系统有髓轴突的节段轴膜上观察到了（Na⁺+K⁺）-ATP酶样免疫反应性。已确定在轴膜的结旁或结间区域未观察到该酶的标记并非由于抗体难以接近这些区域而产生的假象。一些中枢神经系统神经胶质细胞显示出大量的质膜和细胞内（Na⁺+K⁺）-ATP酶样免疫反应性。通过对连续切片中细胞进行神经胶质纤维酸性蛋白免疫反应性标记，在视神经的胞体和放射状突起或小脑中的绒毛状突起中呈阳性，从而将这些细胞鉴定为星形胶质细胞。覆盖在节段轴膜上的星形胶质细胞突起对该酶也呈阳性染色。在少突胶质细胞胞体或其形成髓鞘的突起中未观察到（Na⁺+K⁺）-ATP酶样免疫反应性。相反，有髓施万细胞的质膜在郎飞结区域显示出最强的免疫反应性。虽然在施万细胞与结和结旁相关区域观察到了免疫反应性的局部集中，但在整个施万细胞表面膜上也注意到了较低水平的均匀染色。（摘要截于400字）
