Texture Analysis of Fluorescence Lifetime Images of AT- and GC-rich Regions in Nuclei

We used intensity and fluorescence lifetime microscopy (FLIM) of 3T3 nuclei to investigate the existence of AT-rich and GC-rich regions of the nuclear DNA. Hoechst 33258 (Ho) and 7-aminoactinomycin D (7-AAD) were used as fluorescence probes specific for AT and GC base pairs, respectively. YOYO-1 (Yo...

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Bibliographic Details
Published in:The journal of histochemistry and cytochemistry 2001-11, Vol.49 (11), p.1443-1451
Main Authors: Murata, Shin-ichi, Herman, Petr, Lakowicz, Joseph R
Format: Article
Language:English
Subjects:
3T3 Cells
Animals
AT Rich Sequence
Benzoxazoles
Bisbenzimidazole
Cell Cycle
Cell Nucleus - metabolism
Cell Nucleus - ultrastructure
Dactinomycin - analogs & derivatives
DNA - chemistry
DNA - metabolism
Fluorescent Dyes
GC Rich Sequence
Image Cytometry
Mice
Microscopy, Fluorescence
Quinolinium Compounds
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Summary:We used intensity and fluorescence lifetime microscopy (FLIM) of 3T3 nuclei to investigate the existence of AT-rich and GC-rich regions of the nuclear DNA. Hoechst 33258 (Ho) and 7-aminoactinomycin D (7-AAD) were used as fluorescence probes specific for AT and GC base pairs, respectively. YOYO-1 (Yo) was used as a dye that displays distinct fluorescence lifetimes when bound to AT or GC base pairs. We combined fluorescence imaging of Ho and 7-AAD with time-resolved measurements of Yo and took advantage of an additional information content of the time-resolved fluorescence. Because a single nucleus could not be stained and measured with all three dyes, we used texture analysis to compare the spatial distribution of AT-rich and GC-rich DNA in 100 nuclei in different phases of the cell cycle. The fluorescence intensity-based analysis of Ho- or 7-AAD-stained images indicates increased number and larger size of the DNA condensation centers in the G2/M-phases compared to G0/1-phases. The lifetime-based study of Yo-stained images suggests spatial separation of the AT- or GC-rich DNA regions in the G2/M-phase. Texture analysis of fluorescence intensity and lifetime images was used to quantitatively study the spatial change of condensation and separation of AT- and GC-rich DNA during the cell cycle.
ISSN:0022-1554
1551-5044
DOI:10.1177/002215540104901112