Decay, fluorescence

The attachment of pyrene or another fluorescent marker to a phospholipid or its addition to an insoluble monolayer facilitates their study via fluorescence spectroscopy [163]. Pyrene is often chosen due to its high quantum yield and spectroscopic sensitivity to the polarity of the local environment. In addition, one of several amphiphilic quenching molecules allows measurement of the pyrene lateral diffusion in the mono-layer via the change in the fluorescence decay due to the bimolecular quenching reaction [164,165]. 

For fluorescent compounds and for times in die range of a tenth of a nanosecond to a hundred microseconds, two very successftd teclmiques have been used. One is die phase-shift teclmique. In this method the fluorescence is excited by light whose intensity is modulated sinusoidally at a frequency / chosen so its period is not too different from die expected lifetime. The fluorescent light is then also modulated at the same frequency but with a time delay. If the fluorescence decays exponentially, its phase is shifted by an angle A([) which is related to the mean life, i, of the excited state. The relationship is... 

Figure Cl.5.11. Far-field fluorescence images (A and D), corresponding fluorescence spectra (B and E), and fluorescence decays (C and F) for two different molecules of a carbocyanine dye at a PMMA-air interface.

Figure C1.5.12.(A) Fluorescence decay of a single molecule of cresyl violet on an indium tin oxide (ITO) surface measured by time-correlated single photon counting. The solid line is tire fitted decay, a single exponential of 480 5 ps convolved witli tire instmment response function of 160 ps fwiim. The decay, which is considerably faster tlian tire natural fluorescence lifetime of cresyl violet, is due to electron transfer from tire excited cresyl violet (D ) to tire conduction band or energetically accessible surface electronic states of ITO. (B) Distribution of lifetimes for 40 different single molecules showing a broad distribution of electron transfer rates. Reprinted witli pennission from Lu andXie [1381. Copyright 1997 American Chemical Society.

Keiier R A 1998 Singie-moiecuie identification in flowing sampie streams by fluorescence burst size and intraburst fluorescence decay rate Anal. Chem. 70 1444-51... 

Enderiein J, Goodwin P M, Van Orden A, Ambrose W P, Erdmann R and Keller R A 1997 A maximum likelihood estimator to distinguish single molecules by their fluorescence decays Chem. Phys. Lett. 270 464-70... 

Birch D J S and Imhof R E 1977 A single-photon counting fluorescence decay-time spectrometer J. Phys. E Sol. Instrum. 10 1044-9... 

J/cm2. As previously reported (1), excitation at 532 nm resulted in a shorter lifetime (86 4 microseconds) even at the lowest fluence used (0.1 J/cm2), where, with extensive signal averaging, a final signal-to-noise ratio of about 20 was obtained. As noted above the observed fluorescence decays at 532 nm became increasing non-exponential with increasing laser fluence. 

The significantly faster PuF6(g) fluorescence decay rate found using 532 nm excitation is unlikely to be due to a thermally induced effect (e.g. pyrolysis). The optical absorption coefficients of PuF6(g) at 532 nm is at most twice as large as at 1064 nm (15) Assuming the 1064 nm absorption... 

Figure 4. Energy diagram for 532 nm excitation of PuF g). The 5f electron states of PuF are shown at the left. The solid arrows indicate photon absorption or emission processes. The wavy arrows indicate nonradiative processes by which excited states of PuFg may be lost. The laser-fluence dependent fluorescence decay found at this excitation wavelength can be explained in terms of a bimolecular reaction between PuFg(g) in its 4550 cm l state and PuF (g) to form PuFj(g). It is assumed that PuF (g) is formed via dissociation of the initially populated PuF state.

Table V. Materials Used as Standards for Fluorescence Decay Time, r...

For single exponential fluorescence decay, as is expected for a sample containing just one fluorophore, either the phase shift or the demodulation can be used to calculate the fluorescence lifetime t. When the excitation light is modulated at an angular frequency (o = 2itv, the phase angle f, by which the emission modulation is shifted from the excitation modulation, is related to the fluorescence lifetime by ... 

Fig. 21.8 Room-temperature fluorescent decay profile for Ba3BP30- 2.

Figure 21.23 exhibits the room-temperature fluorescence decay profiles of Ba3BP30i2 Eu powders. The experimental decay curve can be fitted by an equation with two exponential terms corresponding to two decay times of 20 ns (98.97%) and 522 ns (1.03%), respectively. 

With the development of new instrumental techniques, much new information on the size and shape of aqueous micelles has become available. The inceptive description of the micelle as a spherical agglomerate of 20-100 monomers, 12-30 in radius (JJ, with a liquid hydrocarbon interior, has been considerably refined in recent years by spectroscopic (e.g. nmr, fluorescence decay, quasielastic light-scattering), hydrodynamic (e.g. viscometry, centrifugation) and classical light-scattering and osmometry studies. From these investigations have developed plausible descriptions of the thermodynamic and kinetic states of micellar micro-environments, as well as an appreciation of the plurality of micelle size and shape. 

Kuhn et al. observed the fluorescence enhancement and fluorescence decay rate of a single terrylene molecule when a spherical gold nanoparticle was approached to the... 

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-... 

FIG. 12 Simulation of fluorescent decays for dye species located in the aqueous phase following laser pulses in TIR from the water-DCE interface according to Eq. (38). A fast rate constant of excited state decay (10 s ) was assumed in (a). The results showed no difference between infinitely fast or slow kinetics of quenching. On the other hand, a much slower rate of decay can be observed for other sensitizers like Eu and porphyrin species. Under these conditions, heterogeneous quenching associated with the species Q can be readily observed as depicted in (b). (Reprinted with permission from Ref 127. Copyright 1997 American Chemical Society.)... 

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

Homogeneous Time Resolved Fluorescence (HTRF) (Cisbio International) is an assay based on the proximity of a lanthanide cryptate donor and a fluorescent acceptor molecule whose excitation wavelength overlaps that of the cryptate s emission. The utility of this technique is based on the time resolved fluorescence properties of lanthanides. Lanthanides are unique in the increased lifetime of their fluorescence decay relative to other atoms, so a delay in collection of the emission intensity removes the background from other fluorescent molecules. An example of the HTRF assay is a generic protein-protein interaction assay shown in Fig. 2. 

There are, however, problems associated with this method of determining fluorescence lifetimes. First, the phase method is not generally applicable for nonexponential signals and, as we shall see later, there are many cases where the observed fluorescence decay is indeed nonexponential. Second, the method... 

A linear plot indicates that the luminescence decay is exponential. The slope of the line gives kt, and rt can be calculated as above. The lifetime obtained by measuring the decay of P-type delayed fluorescence is equal to one-half the lifetime of the triplet state (see Section 5.2). Since in fluid solution at room temperature phosphorescence is generally much weaker than delayed fluorescence, the measurement of delayed fluorescence decay offers a convenient method for determining the lifetime of triplets at room temperature. 

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