Fluorescence polarization anisotropy properties

Although the structural differences between the water/CClq and water/DCE interfaces are not so large, the chemical and/or physical nature of the organic phase itself reflects on the photophysical properties of a probe molecule, indicating the novelty of the present experimental approaches. Systematic investigations are important to reveal factors governing structural and physical characteristics of water/oil interfaces. Therefore, we introduced fluorescence dynamic anisotropy and excitation energy transfer measurements to other water/oil interfacial systems the data are summarized in Table 12.3. The results are discussed in terms of the relationship between the interfacial stracture and the polarity at the water/oil interfaces (Section 12.6). 

The polarization characteristics of monochromators have important consequences in the measurement of fluorescence anisotropy. Such measurements must be corrected for the varying efficiencies of each optical component this correction is expressed as the G-factor (Section 10.4). However, the extreme properties of the concave gratings (Figure 2.11) can cause difficulties in the measurement of fluorescence polarization. For example, assume that the polarization is to be measured at an excitation wavelength of 450 nm. The excitation intensities will be nearly equal with the excitation polarizers in each orientation, which makes it easier to compare the relative emission intensities. If the nission is unpdarized, the relative intensities of the parallel (U) and perpendicular (X) excit ion will be nearly equal. However, suppose the excic ion is at 340 nm, in which case the intensities the... 

The kinetic interactions between solvent and solute molecules in free solution determine their rotational and translational diffusion characteristics. Fluorescence polarization is a spectroscopic technique that allows the determination of motional preferences of reporter molecules in fluids with respect to both the rate of motion and the orientational restriction of that motion [1,2], For spherical molecules in isotropic fluids at low concentrations, these motions can be described by the Stokes-Einstein and Perrin relationships, and if these motions have an equal probability of occurring in any dimension they are referred to as isotropic. However, when a fluid displays structure, or anisotropy, the motion of diffusing molecules may be restricted, generally to different extents in different dimensions, and these motions are said to be anisotropic. New approaches must then be taken in order to describe the probe s hydrodynamic behavior. By measuring the hydrodynamic properties of a fluorescent probe in solution, it is possible to extract valuable information on the physical structure and properties of a fluid. Knowledge of the physical structure and properties of food fluids and matrices is essential for solving practical problems in food research. 

Abstract The response signal of an immense number of fluorescence reporters with a broad variety of structures and properties can be realized through the observation in changes of a very limited number of fluorescence parameters. They are the variations in intensity, anisotropy (or polarization), lifetime, and the spectral changes that allow wavelength-ratiometric detection. Here, these detection methods are overviewed, and specific demands addressed to fluorescence emitters for optimization of their response are discussed. 

Quenching studies of protein fluorescence provide answers regarding the accessibility of certain internal or external groups to quencher molecules. Another application concerns the study of associative behavior and properties of proteins and membranes. The rationale is that the fluorescence transition is polarized and this polarization can be exploited in time-resolved analysis and interpreted in terms of the rotation or tumbling motion which in turn is determined by the viscosity and structure of the environment of the fluorescing group. In particular, anisotropy decay studies have yielded a great deal of information on the mobility of natural and artificial membranes and/or the dynamics of proteins as well as small molecules in membranes. For such studies fluorescence lifetime labels that can be attached to proteins or that dissolve in membranes have... 

As a further com dicating factor, tryptophan dis days complex spectral properties due to the presence of two nearly isoenergetic excited states. and The electronic transitions display distinct absorption, emission, and anisotropy spectra and are differently sensitive to solvent polarity. The complexity of indole photc hysics has stimulated detailed studies of protein fluorescence but has also inhibited interpretation of the data. 

One of the easiest and most commonly used techniques to study the anisotropic optical properties of dye molecules is fluorescence anisotropy (FA). A dilute solution of the dye is excited with vertically polarized light and the fluorescence emission polarized parallel (I ) and perpendicular (I ) to the electric field vector of the exciting beam are detected with a 90 -scattering geometry, see figure 1. The FA is defined as ... 

A series of aminostilbenes (lA-C, 2A-C) have been synthesized to test the effect of substitution of the amino group upon the photophysics and photochemistry of stilbenes [45]. This study indicated that the photophysics properties of trans-2-aminostilbene, lA, and trans-3-aminostilbene, IB, were similar. as-2-Aminostilbene, 2A, and ds-3-aminostilbene, 2B, showed similar fluorescent lifetimes. Anomalous behavior of emission anisotropy for short-living derivatives of stilbenes was described [46]. The absorption, fluorescence and polarization excitation, and emission spectra... 

Fluorescence anisotropy is one of the properties which can be used in the study of biomolecules and their association. Huorescence anisotropy (r) which can be measured on normal spectrofluorimeters and microplate readers using a polarizing filter in the excitation and another in the emission beam is defined as... 

Emission spectra and fluorescence lifetimes of individual molecules measured in single-molecule experiments often vary substantially from molecule to molecule, and fluctuate on a broad range of time scales [186-188, 191, 192, 201, 232-234]. These fluctuations can reflect varying interactions of the fluorescing molecules with their surroundings, or chemical processes that change the spectroscopic properties, such as cis-trans isomerizations and transitions between oxidized, reduced or triplet states. Emitting molecules sometimes abmptly go dark, presumably as a result of irreversible photochemical reactions. In some cases, anisotropy measurements have shown that the fluctuations in the fluorescence yield are not due simply to rotational reorientation, because the fluorescence remains polarized parallel to the excitation. 

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