Smyrnis E, Kim S U, Kim M W, Oger J, Sylvester C, Paty D W
J Neuroimmunol. 1986 Nov;13(1):47-60. doi: 10.1016/0165-5728(86)90049-4.
We describe here the methods by which enriched populations of oligodendrocytes were isolated from adult porcine brains using Percoll density gradient centrifugation and their immunological properties analyzed by a fluorescence-activated cell sorter (FACS). The analyses by immunofluorescence microscopy and FACS indicated 95% and 93% purity of oligodendrocytes in the cell preparations when galactocerebroside antibody, an oligodendrocyte-specific marker, was employed. Further FACS analysis indicated that 90% of the cells with the forward angle light scatter characteristics of oligodendrocytes were immunolabelled by the monoclonal antibody HNK-1. The FACS sorting capability was then used to correlate the forward angle light scatter and propidium iodide uptake characteristics of our cell preparations to cell debris, erythrocytes as well as viable and non-viable oligodendrocytes. By gating on a multiparameter basis on both defined forward light scatter and positive galactocerebroside immunolabelling, oligodendrocyte populations exceeding 98% purity were obtained which were suitable for long-term culture and further immunological and neurobiological studies.
我们在此描述了从成年猪脑中使用Percoll密度梯度离心法分离富集少突胶质细胞群体的方法，并通过荧光激活细胞分选仪（FACS）分析其免疫学特性。当使用少突胶质细胞特异性标志物半乳糖脑苷脂抗体时，免疫荧光显微镜分析和FACS分析表明细胞制剂中少突胶质细胞的纯度分别为95%和93%。进一步的FACS分析表明，具有少突胶质细胞前向角光散射特征的细胞中有90%被单克隆抗体HNK-1免疫标记。然后利用FACS分选能力将我们细胞制剂的前向角光散射和碘化丙啶摄取特征与细胞碎片、红细胞以及存活和非存活的少突胶质细胞相关联。通过基于定义的前向光散射和阳性半乳糖脑苷脂免疫标记进行多参数门控，获得了纯度超过98%的少突胶质细胞群体，这些群体适合长期培养以及进一步的免疫学和神经生物学研究。
