Bloom G S, Schoenfeld T A, Vallee R B
J Cell Biol. 1984 Jan;98(1):320-30. doi: 10.1083/jcb.98.1.320.
We prepared a monoclonal antibody to microtubule-associated protein 1 (MAP 1), one of the two major high molecular weight MAP found in microtubules isolated from brain tissue. We found that MAP 1 can be resolved by SDS PAGE into three electrophoretic bands, which we have designated MAP 1A, MAP 1B, and MAP 1C in order of increasing electrophoretic mobility. Our antibody recognized exclusively MAP 1A, the most abundant and largest MAP 1 polypeptide. To determine the distribution of MAP 1A in nervous system tissues and cells, we examined tissue sections from rat brain and spinal cord, as well as primary cultures of newborn rat brain by immunofluorescence microscopy. Anti-MAP 1A stained white matter and gray matter regions, while a polyclonal anti-MAP 2 antibody previously prepared in this laboratory stained only gray matter. This confirmed our earlier biochemical results, which indicated that MAP 1 is more uniformly distributed in brain tissue than MAP 2 (Vallee, R.B., 1982, J. Cell Biol., 92:435-442). To determine the identity of cells and cellular processes immunoreactive with anti-MAP 1A, we examined a variety of brain and spinal cord regions. Fibrous staining of white matter by anti-MAP 1A was generally observed. This was due in part to immunoreactivity of axons, as judged by examination of axonal fiber tracts in the cerebral cortex and of large myelinated axons in the spinal cord and in spinal nerve roots. Cells with the morphology of oligodendrocytes were brightly labeled in white matter. Intense staining of Purkinje cell dendrites in the cerebellar cortex and of the apical dendrites of pyramidal cells in the cerebral cortex was observed. By double-labeling with antibodies to MAP 1A and MAP 2, the presence of both MAP in identical dendrites and neuronal perikarya was found. In primary brain cell cultures anti-MAP 2 stained predominantly cells of neuronal morphology. In contrast, anti-MAP 1A stained nearly all cells. Included among these were neurons, oligodendrocytes and astrocytes as determined by double-labeling with anti-MAP 1A in combination with antibody to MAP 2, myelin basic protein or glial fibrillary acidic protein, respectively. These results indicate that in contrast to MAP 2, which is specifically enriched in dendrites and perikarya of neurons, MAP 1A is widely distributed in the nervous system.
我们制备了一种针对微管相关蛋白1(MAP 1)的单克隆抗体,MAP 1是从脑组织分离出的微管中发现的两种主要高分子量微管相关蛋白之一。我们发现,MAP 1经十二烷基硫酸钠聚丙烯酰胺凝胶电泳(SDS PAGE)可分离为三条电泳带,按照电泳迁移率递增的顺序,我们将其分别命名为MAP 1A、MAP 1B和MAP 1C。我们的抗体仅识别MAP 1A,它是最丰富且最大的MAP 1多肽。为了确定MAP 1A在神经系统组织和细胞中的分布,我们通过免疫荧光显微镜检查了大鼠脑和脊髓的组织切片,以及新生大鼠脑的原代培养物。抗MAP 1A抗体对白质和灰质区域均有染色,而本实验室先前制备的一种抗MAP 2多克隆抗体仅对灰质有染色。这证实了我们早期的生化结果,即MAP 1在脑组织中的分布比MAP 2更均匀(瓦利,R.B.,1982年,《细胞生物学杂志》,92:435 - 442)。为了确定与抗MAP 1A发生免疫反应的细胞和细胞突起的身份,我们检查了多种脑和脊髓区域。通常观察到抗MAP 1A对白质的纤维状染色。部分原因是轴突的免疫反应性,这是通过检查大脑皮质中的轴突纤维束以及脊髓和脊神经根中的大型有髓轴突判断得出的。白质中具有少突胶质细胞形态的细胞被强烈标记。在小脑皮质中观察到浦肯野细胞树突以及大脑皮质中锥体细胞顶树突的强烈染色。通过用抗MAP 1A和抗MAP 2抗体进行双重标记,发现相同的树突和神经元胞体中同时存在这两种微管相关蛋白。在原代脑细胞培养物中,抗MAP 2主要对具有神经元形态的细胞染色。相比之下,抗MAP 1A几乎对所有细胞都有染色。通过分别用抗MAP 1A与抗MAP 2、髓鞘碱性蛋白或胶质纤维酸性蛋白的抗体进行双重标记确定,这些细胞包括神经元、少突胶质细胞和星形胶质细胞。这些结果表明,与特异性富集于神经元树突和胞体中的MAP 2不同,MAP 1A在神经系统中广泛分布。