Fluorescence decay analysis

Eaton D. F. (1990) Recommended Methods for Fluorescence Decay Analysis, Pure Appl. Chem. 62, 1631-1648. 

The important theoretical, calculated, and measured quantities for hiexponential fluorescence decay analysis are collected in Table 1. 

Non-linear least-squares fitting by the Marquardt method [19,20] appears to be the most commonly used technique for hiexponential fluorescence decay analysis, at least for a time-domain measurement such as used here [21,22]. Fitting by this method requires evaluation of the derivatives of the model equation (Equation... 

The book does not include data analysis. Analysis of multispectral TCSPC data, diffuse optical tomography data, or fluorescence correlation data differs considerably from traditional fluorescence decay analysis, and many analysis problems are not entirely solved yet. The author believes that data analysis should be the subject of a different book, and leaves this task to someone who is more familiar with it. 

A map of the photoisomerization potential energy surface for tetraphenylethylene in alkane solvents was prepared using a fluorescence and picosecond optical calorimetry (Figure 3.4) [21]. Line shapes of the vertical and relaxed exdted-state emissions at 294 K in methylcyclohexane were obtained from the steady-state emission spec-tmm, the wavelength dependence of the time-resolved fluorescence decays, the temperature dependences of the vertical and relaxed state emission quantum yields, and of the time-resolved fluorescence decays. Analysis of these data in conjunction with values of the twisted exdted-state energy provided values for the energies of the vertical, conformationally relaxed, and twisted exdted states on the photoisomerization surface, as well as the barriers to their interconversion. The energy difference between the last two states is found to be 1.76 0.15 kcal/mol in methylcyclohexane. 

Zachariasse KA, Duveneck G, Kiihnle W et al (1991) Multicomponent fluorescence decay analysis in intramolecular excimer formation with dipyrenylalkanes. In Honda K (ed) Photophysical processes in organized molecular systems. Elsevier, Amsterdam, pp 83... 

The above analysis was entirely consistent with the experimental behavior observed for the systems Eu(III) and anthracene as well as Ru(bpy)3+ and TCNQ at the water-DCE interface [127]. The dependence of the rate of fluorescent decay for Eu(III) on the anthra-... 

Beecham, J. M. Brand, L. Global analysis of fluorescence decay applications to some unusual experimental and theoretical studies. Photochem. Photobiol. 1986, 44, 323-329. 

FRET applications employing CFP and YFP are complicated due to considerable bleed-through between CFP and YFP fluorescence (Figs. 5.5B and 5.6B). Direct excitation of YFP and bleed-through of CFP fluorescence into the YFP detection channel have to be corrected for as shown in Chapters 7 and 8. The multiexponential fluorescence decay of all CFP variants complicates the quantification of FRET by donor lifetime methods. Altogether these factors make quantitative analysis of the FRET efficiency relatively difficult. 

Tleugabulova, D., Duft, A.M., Zhang, Z., Chen, Y., Brook, M.A. and Brennan, J.D. (2004) Evaluating formation and growth mechanisms of silica particles using fluorescence anisotropy decay analysis. Langmuir, 20, 5924—5932. 

We wish to use the fluorescence decay data in Figure 2 to construct the SCP as it propagates through the PMMA film, focusing on data at the final phase of the dissolution process. This analysis will involve the following three assumptions ... 

To answer the question as to whether the fluorescence decay consists of a few distinct exponentials or should be interpreted in terms of a continuous distribution, it is advantageous to use an approach without a priori assumption of the shape of the distribution. In particular, the maximum entropy method (MEM) is capable of handling both continuous and discrete lifetime distributions in a single analysis of data obtained from pulse fluorometry or phase-modulation fluorometry (Brochon, 1994) (see Box 6.1). 

It is interesting to note that when using two fluorophores (whose fluorescence decays are known to be single exponentials), one as a sample and the other as a reference, it is possible to determine the lifetimes of both the fluorophores without external reference this can be achieved in data analysis by varying the reference lifetime until a minimum value of x is reached. 

Beechem f. M., Ameloot M. and Brand L. (1985) Global Analysis of Fluorescence Decay Surfaces Excited-State Reactions, Chem. Phys. Lett. 120, 466-472. 

G(t) decays with correlation time because the fluctuation is more and more uncorrelated as the temporal separation increases. The rate and shape of the temporal decay of G(t) depend on the transport and/or kinetic processes that are responsible for fluctuations in fluorescence intensity. Analysis of G(z) thus yields information on translational diffusion, flow, rotational mobility and chemical kinetics. When translational diffusion is the cause of the fluctuations, the phenomenon depends on the excitation volume, which in turn depends on the objective magnification. The larger the volume, the longer the diffusion time, i.e. the residence time of the fluorophore in the excitation volume. On the contrary, the fluctuations are not volume-dependent in the case of chemical processes or rotational diffusion (Figure 11.10). Chemical reactions can be studied only when the involved fluorescent species have different fluorescence quantum yields. 

M. M. H. Khalil, N. Boens, M. VanDer Auweraer, M. Ameloot, R. Andriessen, J. Hofkens, andF. C. De Schryver, Compartimental analysis of the fluorescence decay surface of the exciplex formation between 1-methylpyrene and triethylamine, J. Phys. Chem. 95,9375-9381 (1991). 

Some methods of quantitative analysis of nonexponential fluorescence decay curves will be shortly described in Section 6.6. 

J. R. Lakowicz, G. Laczko, H. Cherek, E. Gratton, and M. Limkeman, Analysis of fluorescence decay kinetics from variable-frequency phase shift and modulation data, Biophys. J. 46, 463—+77 (1984). 

Single-photon silicon APDs possess a quantum efficiency ofca. 20-40% between 700 and 900 nm which compares very favorably with ca. 3% at best expected from an S20R or SI photocathode over this range. The lack of late-pulsing in an APD response as compared with a linear focused photomultiplier also has some virtues in the reconvolution analysis of fluorescence decay curves. 

W. R Laws and L. Brand, Analysis of two-state excited-state reactions. The fluorescence decay of 2-naphthol, J. Phys. Chem. 83, 795-802 (1979). 

J. R. Knutson, J. M. Beechem, and L. Brand, Simultaneous analysis of multiple fluorescence decay curves A global approach, Chem. Phys. Lett. 102, 501-507 (1983). 

J. B. A. Ross, W. R. Laws, J. C. Sutherland, A. Buku, P. G. Katsoyannis, I. L. Schwartz, and H. R. Wyssbrod, Linked-function analysis of fluorescence decay kinetics Resolution of side-chain rotamer populations of a single aromatic amino acid in small polypeptides, Photochem. Photobiol. 44, 365-370 (1986). 

Proteins having one chromophore per molecule are the simplest and most convenient in studies of fluorescence decay kinetics as well as in other spectroscopic studies of proteins. These were historically the first proteins for which the tryptophan fluorescence decay was analyzed. It was natural to expect that, for these proteins at least, the decay curves would be singleexponential. However, a more complex time dependence of the emission was observed. To describe the experimental data for almost all of the proteins studied, it was necessary to use a set of two or more exponents.(2) The decay is single-exponential only in the case of apoazurin.(41) Several authors(41,42) explained the biexponentiality of the decay by the existence of two protein conformers in equilibrium. Such an explanation is difficult to accept without additional analysis, since there are many other mechanisms leading to nonexponential decay and in view of the fact that deconvolution into exponential components is no more than a formal procedure for treatment of nonexponential curves. 

One can expect that the analysis of continuous distributions of electronic excited-state lifetimes will not only provide a higher level of description of fluorescence decay kinetics in proteins but also will allow the physical mechanisms determining the interactions of fluorophores with their environment in protein molecules to be elucidated. Two physical causes for such distributions of lifetimes may be considered ... 

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