Rougon G, Hirsch M R, Hirn M, Guenet J L, Goridis C
Neuroscience. 1983 Oct;10(2):511-20. doi: 10.1016/0306-4522(83)90149-5.
A monoclonal antibody, designated anti-NSP-4 (anti-Neural cell Surface Protein-4), was obtained from a hybridoma generated by fusing rat myeloma cells with splenocytes of a rat immunized with membranes from the cerebella of weaver mutant mice. This antibody reacted with several high-molecular weight polypeptides in extracts prepared from the newborn and adult CNS of wild-type mice. The main NSP-4-reactive bands from neonatal cerebellum and spinal cord migrated with apparent molecular weights of 220,000 and 140,000. Major bands of 160,000 and of 175,000, 160,000 and 140,000 molecular weight were revealed in the adult cerebellum and spinal cord, respectively. Reaction of the antibodies with concanavalin A-binding proteins demonstrated the glycoprotein nature of the antigen. Cell types expressing NSP-4 antigen were determined using indirect immunofluorescence on monolayer cultures of early postnatal mouse cerebellar and dorsal root ganglion cells and on sections of developing and adult mouse cerebellum. In cerebellar cultures, the antibody reacted with the surface membrane of a subpopulation of astrocytes and of a small subset of neurones. In dorsal root ganglion cultures, anti-NSP-4 antibodies were highly specific for a subclass of small neurones. Staining for NSP-4 in sections of adult cerebellum was confined to the granular layer where the antibody seemed to label astroglia. In the developing cerebellum, NSP-4 staining outlined cell bodies of neuroblasts and migrating granule cells in the external granular layer. Post-migratory granule cells and Purkinje cells were negative. As in the adult, the labeled structures in the internal granular layer were probably astrocytes. Our results on the in vivo and in vitro localization of NSP-4 show its expression by subclasses of neurones and astrocytes in the cerebellum and by a subclass of neurones in cultures from the peripheral nervous system. The developmentally-regulated changes in the molecular weight forms of the NSP-4 antigen together with the shift in its cellular localization during cerebellar ontogeny suggest a functional significance for this antigen in developmental processes.
一种单克隆抗体，命名为抗NSP-4（抗神经细胞表面蛋白-4），是从将大鼠骨髓瘤细胞与用韦弗突变小鼠小脑膜免疫的大鼠脾细胞融合产生的杂交瘤中获得的。该抗体与野生型小鼠新生和成年中枢神经系统提取物中的几种高分子量多肽发生反应。来自新生小脑和脊髓的主要NSP-4反应带的表观分子量分别为220,000和140,000。在成年小脑和脊髓中分别显示出分子量为160,000以及175,000、160,000和140,000的主要条带。抗体与伴刀豆球蛋白A结合蛋白的反应证明了抗原的糖蛋白性质。使用间接免疫荧光法对出生后早期小鼠小脑和背根神经节细胞的单层培养物以及发育中和成年小鼠小脑的切片进行检测，以确定表达NSP-4抗原的细胞类型。在小脑培养物中，该抗体与星形胶质细胞亚群和一小部分神经元的表面膜发生反应。在背根神经节培养物中，抗NSP-4抗体对一小类小神经元具有高度特异性。成年小脑切片中NSP-4的染色局限于颗粒层，抗体似乎在此标记星形胶质细胞。在发育中的小脑中，NSP-4染色勾勒出神经母细胞的细胞体以及外颗粒层中迁移的颗粒细胞。迁移后的颗粒细胞和浦肯野细胞呈阴性。与成年时一样，内颗粒层中标记的结构可能是星形胶质细胞。我们关于NSP-4体内和体外定位的结果表明，它在小脑中由神经元和星形胶质细胞亚类表达，在外周神经系统培养物中由神经元亚类表达。NSP-4抗原分子量形式的发育调节变化以及其在小脑个体发育过程中细胞定位的转变表明该抗原在发育过程中具有功能意义。
