Berti Mattera L N, Larocca J N, de Iraldi A P, Pasquini J M, Soto E F
Departamento de Quimica Biológica, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina.
Neurochem Int. 1984;6(1):41-50. doi: 10.1016/0197-0186(84)90024-x.
A method for the isolation of oligodendroglial cells from young and adult whole rat brains, using a Percoll density gradient is presented. The minced tissue, incubated in a balanced salt solution containing 0.1% trypsin is further dissociated by forcing it through nylon screens to 145 and 74 ?m pore size. The crude suspension is then mixed with an isosmotic Percoll solution and centrifuged for 15 min. An in situ generated density gradient allows the separation of five bands, only one of which (Band C) lying between ?1.050 and ?1.062 contains cellular elements. The isolated cells show the typical morphological characteristics of oligodendroglia. A detailed morphological study of the cells isolated from whole brains of 10-, 30- and 120-day old rats is presented for the first time in the literature and immunocytochemical characterization is carried out using specific (antigalactocerebroside) and non specific (anti-glial fibrillary acidic protein) anti-sera. The method is simple and rapid and isosmotic conditions are maintained throughout, resulting in a better preservation of cell integrity. It represents an improvement over the two previous methods described in the literature and will be useful for studying different developmental events (biochemical and morphological) occurring in oligodendroglial cells at early stages of myelin formation.
本文介绍了一种使用Percoll密度梯度从幼年和成年大鼠全脑中分离少突胶质细胞的方法。将切碎的组织在含有0.1%胰蛋白酶的平衡盐溶液中孵育，然后通过孔径为145和74μm的尼龙筛进一步解离。然后将粗悬液与等渗Percoll溶液混合并离心15分钟。原位生成的密度梯度允许分离出五条带，其中只有一条带（带C）位于1.050至1.062之间，包含细胞成分。分离出的细胞显示出少突胶质细胞的典型形态特征。本文首次在文献中对从10日龄、30日龄和120日龄大鼠全脑中分离出的细胞进行了详细的形态学研究，并使用特异性（抗半乳糖脑苷脂）和非特异性（抗胶质纤维酸性蛋白）抗血清进行了免疫细胞化学表征。该方法简单快速，且始终保持等渗条件，从而更好地保存细胞完整性。它代表了对文献中先前描述的两种方法的改进，将有助于研究髓鞘形成早期少突胶质细胞中发生的不同发育事件（生化和形态学）。
