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Combined light and fluorescent microscopical imaging of nucleolar organizer regions and cellular rRNA as detected by fluorescent in situ hybridization



Combined light and fluorescent microscopical imaging of nucleolar organizer regions and cellular rRNA as detected by fluorescent in situ hybridization



Cytometry 19(2): 171-176



A method for combined light and fluorescent microscopic imaging of nucleolar organizer regions and cellular rRNA is described. Nucleolar organizer regions were detected by silver staining (Ag-NOR), and rRNA was detected by fluorescent in situ hybridization (FISH). MG-63 human fibrosarcoma cells were silver stained prior to in situ hybridization. To quantitate Ag-NOR within individual cells, brightfield images were digitized, and the total Ag-NOR area/nucleus was determined. Fluorescent images were digitized, and the total cellular fluorescence was calculated after correction for nonuniformity of illumination. By using this method, it was shown that the Ag-NOR procedure did not significantly affect the fluorescence intensity related to FISH. Furthermore, the hybridization procedure did not interfere with quantitation of Ag-NOR. With this method, both Ag-NOR and rRNA product can be quantitated within the same cell. Because the relationship of rRNA content to cell proliferation is well established, correlation to quantitative Ag-NOR parameters within individual cells will contribute to the better definition of the relationship of quantitative Ag-NOR indices with cellular proliferation.

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Accession: 008345213

Download citation: RISBibTeXText

PMID: 7743896

DOI: 10.1002/cyto.990190212



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