Open AccessTexture analysis of fluorescence lifetime images of nuclear DNA with effect of fluorescence resonance energy transfer
Author(s) -
Murata Shinichi,
Herman Petr,
Lakowicz Joseph R.
Publication year - 2001
Publication title -
cytometry
Language(s) - English
Resource type - Journals
eISSN - 1097-0320
pISSN - 0196-4763
DOI - 10.1002/1097-0320(20010201)43:2<94::aid-cyto1023>3.0.co;2-4
Subject(s) - fluorescence , förster resonance energy transfer , fluorescence microscope , fluorescence lifetime imaging microscopy , microscopy , texture (cosmology) , phase (matter) , biophysics , nuclear magnetic resonance , chemistry , materials science , optics , biology , physics , artificial intelligence , computer science , image (mathematics) , organic chemistry
Background Fluorescence lifetime imaging microscopy (FLIM) is becoming an important tool in cellular imaging. In FLIM, the image contrast is concentration insensitive, whereas it is sensitive to the local environment and interactions of fluorophores such as fluorescence resonance energy transfer (RET). Methods Fluorescence microscopy, lifetime imaging, and texture analysis were used to study the spatial distribution of fluorophores bound to nuclear DNA. 3T3‐Swiss albino mice fibroblast nuclei were labeled with Hoechst 33258 (Ho), an AT‐specific dye, and 7‐aminoactinomycin D (7‐AAD), a GC‐specific dye. Ho is a RET donor to the 7‐AAD acceptor. Results Texture analysis of 50 alcohol‐fixed nuclei quantitatively showed changes of spatial distribution of apparent donor lifetimes. RET increased the spatial heterogeneity in the phase and modulation lifetime images. In most of the doubly stained cells (about 80%), the phase and modulation lifetime distributions were spatially homogeneous. In about 20% of the cells, we noticed that lower phase and modulation lifetimes caused by RET were correlated with regions of high Ho intensity in the nuclei. Conclusions The spatial lifetime heterogeneity of Ho in presence of 7‐AAD seems to be caused by RET between closely spaced strands in the three dimensionally condensed regions of DNA. Cytometry 43:94–100, 2001. © 2001 Wiley‐Liss, Inc.