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Time-averaged fluorescence intensity analysis in fluorescence fluctuation polarization sensitive experiments.

Turgeman L, Fixler D - Biomed Opt Express (2013)

Bottom Line: In this paper, the variance of the time-averaged fluorescence intensity extracted from the second moment of the measured fluorescence intensity is analyzed in the short time limit, before fluctuations resulting from rotational diffusion average out.Since rotational correlation times of fluorescence molecules are typically much lower than the temporal resolution of the system, independently of the time bins used, averaging over an ensemble of time-averaged trajectories was performed in order to construct the time-averaged intensity distribution, thus improving the signal-to-noise ratio.Rotational correlation times of fluorescein molecules in different viscosities of the medium within the range of the anti-bunching time (1-10 ns) were then extracted using this method.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Engineering and Institute of Nanotechnology and Advanced Materials, Bar Ilan University, Ramat Gan, 52900, Israel ; Tur.Lior@gmail.com.

ABSTRACT
In fluorescence fluctuation polarization sensitive experiments, the limitations associated with detecting the rotational timescale are usually eliminated by applying fluorescence correlation spectroscopy analysis. In this paper, the variance of the time-averaged fluorescence intensity extracted from the second moment of the measured fluorescence intensity is analyzed in the short time limit, before fluctuations resulting from rotational diffusion average out. Since rotational correlation times of fluorescence molecules are typically much lower than the temporal resolution of the system, independently of the time bins used, averaging over an ensemble of time-averaged trajectories was performed in order to construct the time-averaged intensity distribution, thus improving the signal-to-noise ratio. Rotational correlation times of fluorescein molecules in different viscosities of the medium within the range of the anti-bunching time (1-10 ns) were then extracted using this method.

No MeSH data available.


Related in: MedlinePlus

The rotational correlation time τr measured from the fluorescence fluctuation measurements of 1 nM fluorescein solution (blue rhombus), and from the bulk time-resolved anisotropy decay measurements for 1 µM fluorescein solution (red squares), versus viscosity. The solid straight line fits the fluorescence fluctuation measurements and the dashed line fits to bulk time-resolved anisotropy decay measurements obtained with the same set-up.
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g008: The rotational correlation time τr measured from the fluorescence fluctuation measurements of 1 nM fluorescein solution (blue rhombus), and from the bulk time-resolved anisotropy decay measurements for 1 µM fluorescein solution (red squares), versus viscosity. The solid straight line fits the fluorescence fluctuation measurements and the dashed line fits to bulk time-resolved anisotropy decay measurements obtained with the same set-up.

Mentions: (a) and (b), for various glycerol percentages (0% — black, 20% — red, 60% — blue, 80% — green). Theory (dotted lines) and simulation (thick solid lines) are compared to experimental results (thin solid lines). decreases in time for all samples. However, fluctuations for the less viscous samples are averaged out faster than fluctuations for the more viscous samples.


Time-averaged fluorescence intensity analysis in fluorescence fluctuation polarization sensitive experiments.

Turgeman L, Fixler D - Biomed Opt Express (2013)

The rotational correlation time τr measured from the fluorescence fluctuation measurements of 1 nM fluorescein solution (blue rhombus), and from the bulk time-resolved anisotropy decay measurements for 1 µM fluorescein solution (red squares), versus viscosity. The solid straight line fits the fluorescence fluctuation measurements and the dashed line fits to bulk time-resolved anisotropy decay measurements obtained with the same set-up.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3675866&req=5

g008: The rotational correlation time τr measured from the fluorescence fluctuation measurements of 1 nM fluorescein solution (blue rhombus), and from the bulk time-resolved anisotropy decay measurements for 1 µM fluorescein solution (red squares), versus viscosity. The solid straight line fits the fluorescence fluctuation measurements and the dashed line fits to bulk time-resolved anisotropy decay measurements obtained with the same set-up.
Mentions: (a) and (b), for various glycerol percentages (0% — black, 20% — red, 60% — blue, 80% — green). Theory (dotted lines) and simulation (thick solid lines) are compared to experimental results (thin solid lines). decreases in time for all samples. However, fluctuations for the less viscous samples are averaged out faster than fluctuations for the more viscous samples.

Bottom Line: In this paper, the variance of the time-averaged fluorescence intensity extracted from the second moment of the measured fluorescence intensity is analyzed in the short time limit, before fluctuations resulting from rotational diffusion average out.Since rotational correlation times of fluorescence molecules are typically much lower than the temporal resolution of the system, independently of the time bins used, averaging over an ensemble of time-averaged trajectories was performed in order to construct the time-averaged intensity distribution, thus improving the signal-to-noise ratio.Rotational correlation times of fluorescein molecules in different viscosities of the medium within the range of the anti-bunching time (1-10 ns) were then extracted using this method.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Engineering and Institute of Nanotechnology and Advanced Materials, Bar Ilan University, Ramat Gan, 52900, Israel ; Tur.Lior@gmail.com.

ABSTRACT
In fluorescence fluctuation polarization sensitive experiments, the limitations associated with detecting the rotational timescale are usually eliminated by applying fluorescence correlation spectroscopy analysis. In this paper, the variance of the time-averaged fluorescence intensity extracted from the second moment of the measured fluorescence intensity is analyzed in the short time limit, before fluctuations resulting from rotational diffusion average out. Since rotational correlation times of fluorescence molecules are typically much lower than the temporal resolution of the system, independently of the time bins used, averaging over an ensemble of time-averaged trajectories was performed in order to construct the time-averaged intensity distribution, thus improving the signal-to-noise ratio. Rotational correlation times of fluorescein molecules in different viscosities of the medium within the range of the anti-bunching time (1-10 ns) were then extracted using this method.

No MeSH data available.


Related in: MedlinePlus