Gebicke-Härter P J, Althaus H H, Rittner I, Neuhoff V
J Neurochem. 1984 Feb;42(2):357-68. doi: 10.1111/j.1471-4159.1984.tb02686.x.
A method is described by which oligodendrocytes from adult pig brains can be isolated. It results in a cellular preparation suitable for long-term culture. The entire procedure can be accomplished within 2-3 h. The purity of oligodendrocytes ranges between 80 and 95% depending on the Percoll gradient used and on the time in vitro. Yields between 2.5 and 4 X 10(7) cells per brain and plating efficiencies on the order of 60% make the system very useful for biochemical investigations. It was shown by immunocytochemical studies that oligodendrocytes produce extensive networks of processes, some of them having elaborate membranous expansions. Anti-galactocerebroside (GC) antibodies as well as anti-myelin basic protein (MBP), anti-Wolfgram protein (WP), anti-glial fibrillary acidic protein (GFAP), and monoclonal antibodies O1 and O4 are used to identify the cell types and to characterize the cellular composition of the cultures. Anti-GC and O1 are suitable markers for these oligodendrocytes. Both antibodies label similar cells, and the staining intensities are equally strong. In the case of O4, variable staining intensities are observed, and a few additional cells are labeled that are anti-GC-. After 3 1/2 weeks in culture, about 60% of the cells can be labeled by anti-MBP. Here too differences in staining intensities are observed. The anti-WP stain is too weak to be defined as positive. The percentage of GFAP+ cells lies in the range 15-20% at maximum. Cells were also mixed into collagen gels. This method appears to be more useful for outgrowth and branching of fibers than are monolayer systems. Drawbacks, however, include limited access for the antibodies and poor recovery of undamaged cells with their fibers.
本文描述了一种从成年猪脑分离少突胶质细胞的方法。该方法能得到适合长期培养的细胞制剂。整个过程可在2 - 3小时内完成。少突胶质细胞的纯度在80%至95%之间，这取决于所用的Percoll梯度和体外培养时间。每只脑的产量在2.5×10⁷至4×10⁷个细胞之间，接种效率约为60%，这使得该系统对生化研究非常有用。免疫细胞化学研究表明，少突胶质细胞能产生广泛的突起网络，其中一些具有精细的膜性扩张。抗半乳糖脑苷脂（GC）抗体以及抗髓鞘碱性蛋白（MBP）、抗Wolfgram蛋白（WP）、抗胶质纤维酸性蛋白（GFAP）和单克隆抗体O1和O4被用于鉴定细胞类型和表征培养物的细胞组成。抗GC和O1是这些少突胶质细胞的合适标记物。两种抗体标记的细胞相似，染色强度相同。对于O4，观察到染色强度可变，并且标记了一些额外的抗GC阴性细胞。培养3.5周后，约60%的细胞可被抗MBP标记。这里也观察到染色强度的差异。抗WP染色太弱，不能定义为阳性。GFAP阳性细胞的百分比最高在15% - 20%范围内。细胞也被混入胶原凝胶中。与单层系统相比，这种方法似乎对纤维的生长和分支更有用。然而，缺点包括抗体难以接近细胞以及未受损细胞及其纤维的回收率低。
