Furuya S, Hashikawa T, Irie F, Hasegawa A, Nakao T, Hirabayashi Y
Laboratory for Glyco-Cell Biology, Frontier Research Program, Institute of Physical and Chemical Research (RIKEN), Saitama, Japan.
Neurosci Res. 1995 Jul;22(4):411-21. doi: 10.1016/0168-0102(95)00920-o.
Monoclonal antibodies (mAbs) against GM1b ganglioside were raised by immunizing NZB/n mice with the antigen purified from bovine brains, and the details of binding specificity of the mAbs were characterized. Anti-GM1b mAb, termed NA-6, reacted specifically with GM1b (NeuAc) and GM1b(NeuGc). NA-6 antibody did not react with other structurally related gangliosides, indicating that the antibody recognizes NeuAc or NeuGc alpha 2-3Gal beta 1-4GalNAc beta 1-4Gal structure. Using NA-6 antibody, GM1b ganglioside in developing rat brain was investigated by TLC/enzyme-immunostaining and detected first on gestational day 16. The specific content of brain GM1b increased until postnatal day 10, and then gradually decreased in later stage of development. Immunohistochemically GM1b was found in proximal dendrites and cell bodies of neurons in extensive regions of adult rat brain. The immunoreactivity tended to be confined in patch-like clusters on cell membranes, as typically found in the hippocampus and neocortex. The GM1b synthase activity, when assayed in the cerebellar microsome proteins, was significantly reduced in lurcher mutant mouse that is devoid of both cerebellar granule and Purkinje cells. These findings demonstrate that GM1b ganglioside exists in neurons and is actively synthesized during the development in rat brain.
通过用从牛脑中纯化的抗原免疫NZB/n小鼠，制备了抗GM1b神经节苷脂的单克隆抗体（mAb），并对这些单克隆抗体的结合特异性细节进行了表征。名为NA-6的抗GM1b单克隆抗体与GM1b（NeuAc）和GM1b（NeuGc）特异性反应。NA-6抗体不与其他结构相关的神经节苷脂反应，表明该抗体识别NeuAc或NeuGcα2-3Galβ1-4GalNAcβ1-4Gal结构。使用NA-6抗体，通过TLC/酶免疫染色研究了发育中大鼠脑中的GM1b神经节苷脂，并在妊娠第16天首次检测到。脑GM1b的特定含量在出生后第10天之前增加，然后在发育后期逐渐下降。免疫组织化学研究发现，GM1b存在于成年大鼠脑广泛区域的神经元近端树突和细胞体中。免疫反应性倾向于局限在细胞膜上的斑块状簇中，如在海马体和新皮质中常见的那样。当在缺乏小脑颗粒细胞和浦肯野细胞的蹒跚突变小鼠的小脑微粒体蛋白中检测时，GM1b合酶活性显著降低。这些发现表明，GM1b神经节苷脂存在于神经元中，并在大鼠脑发育过程中被积极合成。
