Immunofluorescent localisation of cytokeratin antigens in mitotic HeLa cells using monoclonal antibodies
Turner, B.M.; Ruane, M.
European Journal of Cell Biology 36(1): 48-57
1985
ISSN/ISBN: 0171-9335 PMID: 2579815 Document Number: 265099
The organization of cytokeratin filaments in mitotic HeLa cells was analyzed by immunofluorescence microscopy using a monoclonal antibody which recognizes proteins with apparent subunit MW 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. Extensive disaggregation and reformation of cytokeratin filaments may occur during mitosis and disaggregated cytokeratin proteins may frequently be located adjacent to mitotic chromosomes.