Fluorescence lifetime analysis of DNA intercalated ethidium bromide and quenching by free dye

Heller, Dennis; Greenstock, C.L.

doi:10.1016/0301-4622(93)e0101-a

Cited by 183 publications

(88 citation statements)

References 14 publications

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“…Our study did demonstrate changes in the peak excitation wavelength when the dyes were bound to the different substrates. These alterations are postulated to indicate different binding modes of the dyes and changes in the accessibility of the dye molecules to the solvent environment (Heller and Greenstock 1994). Phase-sensitive flow cytometric analysis of the DNAbinding fluorochromes provided fluorescence lifetime values in accord with previously published results (Netzel et al 1995;Hochstrasser and Millar 1992;Wadkins and Jovin 1991;Olmsted and Kearns 1977).…”

Section: Discussionsupporting

confidence: 82%

“…Phase-sensitive flow cytometry provides the capability to quantify fluorescence lifetimes and to resolve the emissions of multiple fluorochrome labels that have different lifetime values (Steinkamp and Crissman 1993). Quantitation of the lifetime of DNAbound fluorochromes provides information on the molecular interactions of a fluorochrome within the target molecule (Sailer et al 1996;Heller and Greenstock 1994). The lifetimes of DNA-bound fluorochromes are altered by physical and chemical factors such as solvents, cations, pH, energy transfer, and quenching.…”

mentioning

confidence: 99%

“…Alterations in chromatin structure change the accessibility of the DNA-bound fluorochromes and influence both the intensity (Evenson et al 1986;Darzynkiewicz et al 1984) and the lifetime of the bound dye (Sailer et al 1996;Heller and Greenstock 1994;Lakowicz 1992;Dewey 1991;Jameson and Reinhart 1989). Differences in the structure of fluorochrome molecules and in their mode of DNA binding lead to differences in D 2 O-induced enhancement in the intensity and lifetime.…”

mentioning

confidence: 99%

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Differential Effects of Deuterium Oxide on the Fluorescence Lifetimes and Intensities of Dyes with Different Modes of Binding to DNA

Sailer

Nastasi

Valdez

et al. 1997

J Histochem Cytochem.

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SUMMARY Deuterium oxide (D 2 O) increases both the fluorescence lifetime and the fluorescence intensity of the intercalating dyes propidium iodide (PI) and ethidium bromide (EB) when bound to nucleic acid structures. We have used spectroscopic analysis coupled with conventional and phase-sensitive flow cytometry to compare the alterations in intensity and lifetime of various DNA-binding fluorochromes bound to DNA and Chinese hamster ovary (CHO) cells in the presence of D 2 O vs phosphate-buffered saline (PBS). Spectroscopic and flow cytometric studies showed a differential enhancement of intensity and lifetime based on the mode of fluorochrome-DNA interaction. The fluorescence properties of intercalating probes, such as 7-aminoactinomycin D (7-AAD) and ethidium homodimer II (EthD II) were enhanced to the greatest degree, followed by the probes TOTO and YOYO, and the non-intercalating probes Hoechst 33342 (HO) and 4 Ј ,6-diamidino-2-phenylindole (DAPI). The non-intercalating probe mithramycin (MI) gave unexpected results, showing a great enhancement of fluorescence intensity and lifetime in D 2 O, indicating that when staining is performed in PBS, much of the MI fluorescence is quenched by the solvent environment. Apoptotic subpopulations of HL-60 cells had a shorter lifetime compared to nonapoptotic subpopulations when stained with EthD II. These results indicate that accessibility of the dye molecules to the solvent environment, once bound to DNA, leads to the differential enhancement effects of D 2 O on fluorescence intensity and lifetime of these probes. D na-binding fluorochromes have been used extensively in many applications, including flow cytometry and gel and capillary electrophoresis. Fluorochromes utilized in these applications must satisfy several criteria. They must bind specifically to nucleic acids, there must be a low quantum yield as free dye in solution, and there must be enhancement of quantum yield on binding to nucleic acid structures. Several classes of DNA-specific fluorochromes with different modes of base pair ( bp ) binding are available: (a) the DNA intercalators propidium iodide (PI) and ethidium bromide (EB), which lack appreciable bp specificity; (b) DNA intercalators with A-T bp preference, such as ethidium homodimer II (EthD II) or with G-C preference, such as 7-amino actinomycin D (7-AAD); and (c) non-intercalating probes with either A-T bp specificity, such as Hoechst 33342 (HO) or 4 Ј ,6-diamidino-2-phenylindole (DAPI) or with G-C specificity, such as mithramycin (MI). Two high quantum yield cyanine fluorochromes, TOTO and YOYO, are dimers of the dyes thiazole orange and oxazole yellow, respectively (Lee et al. 1986). These fluorochromes are believed to bind to DNA by intercalation, without base specificity (Haugland 1992) but with some base sequence preference (Netzel et al. 1995).Enhancement of the fluorescence intensity of DNAbinding probes improves the signal-to-noise ratio, allowing the use of lower dye concentrations and thereby reducing potential self-quenching between d...

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Section: Discussionsupporting

confidence: 82%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Differential Effects of Deuterium Oxide on the Fluorescence Lifetimes and Intensities of Dyes with Different Modes of Binding to DNA

Sailer

Nastasi

Valdez

et al. 1997

J Histochem Cytochem.

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“…6) are 6.07(± 0.18) × 10 4 M − 1 for 3 and 3.72(± 0.10) × 10 4 M − 1 for 8; they may be considered rather high and verify the tight binding of compounds 3 and 8 to DNA [29,32]. Taking τ o = 23 ns as the fluorescence lifetime of EB-DNA system [33], the quenching constants of the compounds as calculated ; the values of the k q constants are significantly higher than 10 10 M −1 s −1 suggesting, thus, the quenching via a static mechanism [34]. A current study from our group, concerning the photo-cleavage of sulfonyl amidoximes [35], show results which are in general agreement with the observations of the present.…”

Section: Dna Binding Studiesmentioning

confidence: 91%

Alkyl and aryl sulfonyl p-pyridine ethanone oximes are efficient DNA photo-cleavage agents

Andreou

Dafnopoulos

Tortopidis

et al. 2016

Journal of Photochemistry and Photobiology B: Biology

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Sulfonyloxyl radicals, readily generated upon UV irradiation of p-pyridine sulfonyl ethanone oxime derivatives, effectively cleave DNA, in a pH independent manner, and under either aerobic or anaerobic conditions. pPyridine sulfonyl ethanone oxime derivatives were synthesized from the reaction of p-pyridine ethanone oxime with the corresponding sulfonyl chlorides in good to excellent yields. All compounds, at a concentration of 100 μM, were irradiated at 312 nm for 15 min, after incubation with supercoiled circular pBluescript KS II DNA and resulted in extended single-and double-strand cleavages. The cleavage ability was found to be concentration dependent, with some derivatives exhibiting activity even at nanomolar levels. Besides that, p-pyridine sulfonyl ethanone oxime derivatives showed good affinity to DNA, as it was observed with UV interaction and viscosity experiments with CT DNA and competitive studies with ethidium bromide. The compounds interact to CT DNA probably by non-classical intercalation (i.e. groove-binding) and at a second step they may intercalate within the DNA base pairs. The fluorescence emission spectra of pre-treated EB-DNA exhibited a significant or moderate quenching. Comparing with the known aryl carbonyloxyl radicals the sulfonyloxyl ones are more powerful, with both aryl and alkyl sulfonyl substituted derivatives to exhibit DNA photo-cleaving ability, in significantly lower concentrations. These properties may serve in the discovery of new leads for "on demand" biotechnological and medical applications.

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“…Throughout the years, a number of fluorimetric methods for the determination of nucleic acids have been developed with ethidium bromide, [40][41][42] lanthanide cations, [43][44][45] ruthenium complexes, [46][47][48] and asymmetric cyanine dyes as fluorescence probes. [49][50][51] Despite the prominence of fullerenes in bionanotechnology, the exploration of their fluorescent properties in solution remains still at a very early age.…”

Section: Introductionmentioning

confidence: 99%

Binding fullerenol C60(OH)24 to dsDNA

Pinteală¹,

Dascalu²,

Ungurenasu³

2009

IJN

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Abstract:The first C 60 (OH) 24 -DNA complex and its fluorescence enhancement is reported. The enhanced fluorescence intensity of fullerenol C 60 (OH) 24 is in proportion to the concentration of DNA in the range of 1 × 10 -9 to 8 × 10 -5 molL -1 and the detection limit was 1.3 ng mL -1 . Fullerenol C 60 (OH) 24 binds significantly to the phosphate backbone of native dsDNA and to base-pairs within the major groove of sodium salt of dsDNA.

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Fluorescence lifetime analysis of DNA intercalated ethidium bromide and quenching by free dye

Cited by 183 publications

References 14 publications

Differential Effects of Deuterium Oxide on the Fluorescence Lifetimes and Intensities of Dyes with Different Modes of Binding to DNA

Differential Effects of Deuterium Oxide on the Fluorescence Lifetimes and Intensities of Dyes with Different Modes of Binding to DNA

Alkyl and aryl sulfonyl p-pyridine ethanone oximes are efficient DNA photo-cleavage agents

Binding fullerenol C60(OH)24 to dsDNA

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