Fluorescence measurements, intensified

Figure 1. Instrumental configuration for fluorescence measurements with an intensified...

In frequency-domain FLIM, the optics and detection system (MCP image intensifier and slow scan CCD camera) are similar to that of time-domain FLIM, except for the light source, which consists of a CW laser and an acousto-optical modulator instead of a pulsed laser. The principle of lifetime measurement is the same as that described in Chapter 6 (Section 6.2.3.1). The phase shift and modulation depth are measured relative to a known fluorescence standard or to scattering of the excitation light. There are two possible modes of detection heterodyne and homodyne detection. 

By means of time-resolved fluorescence studies we were able to determine the C60 fluorescence deactivation rates, as 2.1 x 1010 s-1 in 9a, 6.6 x 109 s 1 in 9b and 1.3 x 109 s-1 in 9c. Importantly, the indulging trend resembles the relationship between the quantum yields of the conjugates (9a-d) and reference (1). In short, an intensified excited-state deactivation emerges with decreasing bridge length. However, no measurable decay rates were found for the trimer 9d. Conclusively, the indirect or direct population of Cgo possibly leads to an exothermic electron-transfer reaction, resulting in the radical-ion-pair state ... 

The peak wavelength of the light emitted from the fluorescent plate of the intensifier is around 520 nm, which matches well the a-Si H photosensitivity spectrum. The image resolution is measured to be 20% better than in conventional intensifier tubes. 

In the past, molecular luminescence spectrometry was always conducted with single channel systems involving a photomultiplier tube (PMT) as the detector. The availability of multichannel detectors with internal gain has provided a new powerful tool for luminescence measurements, and several types of applications have been reported (1-15). This paper is concerned with the application of an intensified diode array dynamic molecular fluorescence and chemiluminescence measurements. In this paper the types of measurements and analytical systems for which multichannel detectors are used in our laboratory are introduced. Next the specific IDA system used is presented along with important hardware and software considerations. Third, the characteristics of the IDA detector are reviewed to give some perspective about its influence on the quality of measurements. Finally, some typical applications to chemical systems are presented to illustrate the advantages of multichannel detection. 

The fluorescence intensity is a measure of alkoxyamine reaction product and it is seen that the emission intensify increases with time without any induction period. At the highest concentration, this is linear and a straight line has been... 

In order to put into evidence the Trp residues thet participate in the emission of the protein, quenching resolved emission with cesium were first performed. When a protein contains two classes of intrin fluoropliore. one at the surface of the protein and the second embedded in the protein matrix, fluorescence intensify quenching with cesium allows obtaining the spectra of these two classes. A selective quenching implies that addition of quencher induces a decrease in the fluorescence observables (intensity, anisotropy and lifetime) of the accessible class. At high quencher concentration the remaining observables measured will reflect essentially those of the embedded fluorophore residues. 

A.B. Thompson, E.M. Sevick-Muracka, Near-infrared fluorescence contrast-enhanced imaging with intensified charge-coupled device homodyne detection measurement precision and accuracy, J. Biomed. Opt. 8, 111-120 (2003)... 

Schlatterer and Schaloske (35) used a low-light SIT camera (Heimann, Wiesbaden, Germany). However, to increase the sensitivity Sonnemann et al. (4) used an intensified CCD camera (model HLA, Proxitronic, Bensheim, Germany) to capture images of the cells. Use of the CCD camera increases the sensitivity of the image captured by a factor of 10. This allows more accurate measurements of cells that have incorporated low amounts of the indicator therefore are only weakly fluorescent. Levels of less than 10 pM of the indicator were able to be detected (4). 

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