Immunofluorescent localisation of cytokeratin antigens in mitotic HeLa cells using monoclonal antibodies.
Immunofluorescent localisation of cytokeratin antigens in mitotic HeLa cells using monoclonal antibodies.
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使用单克隆抗体对有丝分裂 HeLa 细胞中的细胞角蛋白抗原进行免疫荧光定位。
DOI：
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发表时间：
1985
影响因子：
6.6
通讯作者：
M. Ruane
中科院分区：
文献类型：
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作者：
B. M. Turner;M. Ruane
使用单克隆抗体通过免疫荧光显微镜分析了有丝分裂 HeLa 细胞中细胞角蛋白丝的组织，该单克隆抗体可识别表观亚基分子量为 52 kDa 和 57 kDa 的蛋白质，并且仅与细胞角蛋白型丝状体结合。通过低渗膨胀、离心到载玻片上、用 0.1% Triton X-100 短暂预提取并固定在 80% 乙醇中，制备有丝分裂细胞用于显微镜分析。该程序对中间丝提供了特别好的分辨率并保留了染色体形态。在前中期-中期细胞中，抗原存在于完全或部分包围染色体的吻合丝网络中，以丝片段和细胞质聚集体的形式存在。有丝分裂后期或后期的细胞不存在表观染色体丝网络。在这些细胞中，非丝状抗原通常位于限定单个染色体外围的窄带中以及可变数量的细胞质丝或片段中。结果表明，细胞角蛋白丝在有丝分裂期间发生广泛的解聚和重组，并且解聚的细胞角蛋白经常位于有丝分裂染色体附近。
The organisation of cytokeratin filaments in mitotic HeLa cells has been analysed by immunofluorescence microscopy using a monoclonal antibody which recognises proteins with apparent subunit molecular weights of 52 kDa and 57 kDa and which binds exclusively to cytokeratin-type filaments. Mitotic cells were prepared for microscopic analysis by hypotonic swelling, centrifugation onto glass slides, brief pre-extraction with 0.1% Triton X-100 and fixation in 80% ethanol. This procedure gave particularly good resolution of intermediate filaments and preservation of chromosome morphology. In prometaphase-metaphase cells the antigen was present in an anastomosing filament network which completely or partially enclosed the chromosomes, in filament fragments and in cytoplasmic aggregates. The epichromosomal filament network was absent from cells in anaphase or later stages of mitosis. In these cells non-filamentous antigen was often located in a narrow band defining the periphery of individual chromosomes and in variable numbers of cytoplasmic filaments or fragments. The results suggest that extensive disaggregation and reformation of cytokeratin filaments occurs during mitosis and that disaggregated cytokeratin proteins are frequently located adjacent to mitotic chromosomes.