摘要
本文报告对人脑胶质瘤细胞系BT_(325)细胞骨架的免疫荧光研究,并比较了几种固定剂和缓冲液对不同细胞骨架成分的影响。除微管及微丝均被染色外,所有细胞均为波形纤维蛋白(vimentin)染色阳性,但只有部分细胞为胶质纤维酸性蛋白(GFAP)染色阳性。说明vimentin是BT_(325)细胞内中间丝的主要成分,而GFAP仅在部分细胞内表达。实验结果还表明,甲醇与甲醛-丙酮、甲醛-Triton固定微管效果较好,而甲醇、醋酸酒精适用于中间丝的固定,甲醛固定也适用于微丝染色。如用Triton处理后进行染色,则缓冲液成分对微管与中间丝影响较大,而微丝不大受影响。本文还观察到,如在Triton处理后先用甲醛固定,再进行免疫染色,则vimentin染色很弱。如先进行免疫染色,然后再用甲醛固定,则vimentin染色很强。结果表明,甲醛固定对vimentin-分子上的抗原位点起遮蔽或破坏作用。
The cytoskeleton of BT_(325), a human glioma cell line, was studied by immunofluorescence microscopy. The effects of different fixatives and buffers on microtubules, microfilaments and intermediate filaments were also compared. It was observed that besides microtubules and microfilaments all cells expressed vimentin. However, only a small fraction of the cells were GFAP positive. We conclude that vimentin is the main component of the intermediate filaments in BT_(325) cells. It was also observed that methanol fixation and formaldehyde fixation followed by acetone or Triton X-100 treatment gave rise to satisfactory results for microtubule immune-staining, and methanol or acidic alcohol fixation resulted in bright staining of intermediate filaments. Formaldehyde fixation also resulted in excellent staining for mierotubnles, but weaker staining for intermediate filaments.If the cells were immune-stained after treatment with Triton X-100, the composition of the buffers had profound effects on microtubules and intermediate filaments, but less effects on microfilaments.It was also observed that if Triton-treated ceils were fixed with formaldehyde before immunostaining with monoclonal antibodies to vimentin, the staining was very weak. However, if the ceils were immunostained first and then fixed with for-maldehyde the staing was quite bright. We conclude that for-maldehyd e fixation may 'mask' or destroy certain epitopes on vimentin molecules.
出处
《解剖学报》
CAS
CSCD
北大核心
1990年第3期269-273,共5页
Acta Anatomica Sinica
关键词
脑胶质瘤
细胞系BT325
细胞骨架
Human glioma cell line BT_(325)
Cytoskeleton, Immunofluorescence
