Imaging fluorescence spectroscopy

X-ray Photoelectron Spectroscopy XPSj Circular Dichroism Spectroscopy Nuclear Magnetic Resonance NMR Imaging Fluorescence Spectroscopy Phosphorescence Spectroscopy Luminescence Spectroscopy Light Scattering X-ray Diffraction Electron Diffraction Microscopy Thermal - M hanical Methods... 

Sanchez E J, Novotny L, Floltom G R and Xie X S 1997 Room-temperature fluorescence imaging and spectroscopy of single molecules by two-photon excitation J. Chem. Phys. A 101 7019-23... 

A nano-light-source generated on the metallic nano-tip induces a variety of optical phenomena in a nano-volume. Hence, nano-analysis, nano-identification and nanoimaging are achieved by combining the near-field technique with many kinds of spectroscopy. The use of a metallic nano-tip applied to nanoscale spectroscopy, for example, Raman spectroscopy [9], two-photon fluorescence spectroscopy [13] and infrared absorption spectroscopy [14], was reported in 1999. We have incorporated Raman spectroscopy with tip-enhanced near-field microscopy for the direct observation of molecules. In this section, we will give a brief introduction to Raman spectroscopy and demonstrate our experimental nano-Raman spectroscopy and imaging results. Furthermore, we will describe the improvement of spatial resolution... 

Conventional TCSPC equipment has been successfully employed in LSM for fluorescence spectroscopy on discrete microscopic volumes [18, 19] and for fluorescence lifetime imaging at a low acquisition speed [1], The use of conventional TCSPC equipment for imaging results in very long acquisition times, several to many minutes per (time-resolved) image. Importantly, operating the TCSPC detection system at too high detection rates, above 5% of the excitation frequency, results in distortion of the recorded decay curve [20],... 

Hanley, Q. S., Verveer, P. J. and Jovin, T. M. (1999). Spectral imaging in a programmable array microscope by hadamard transform fluorescence spectroscopy. Appl. Spectrosc. 53, 1-10. 

Vukjovic et al.199 recently proposed a simple, fast, sensitive, and low-cost procedure based on solid phase spectrophotometric (SPS) and multicomponent analysis by multiple linear regression (MA) to determine traces of heavy metals in pharmaceuticals. Other spectroscopic techniques employed for high-throughput pharmaceutical analysis include laser-induced breakdown spectroscopy (LIBS),200 201 fluorescence spectroscopy,202 204 diffusive reflectance spectroscopy,205 laser-based nephelometry,206 automated polarized light microscopy,207 and laser diffraction and image analysis.208... 

Lakowicz J. R. and Szymacinski H. (1996) Imaging Applications of Time-Resolved Fluorescence Spectroscopy, in Wang X. F. and Herman B. (Eds), Fluorescence Imaging Spectroscopy and Microscopy, Chemical Analysis Series, Vol. 137, John Wiley ... 

Finally, in Chapter 11 some advanced techniques are briefly described fluorescence up-conversion, fluorescence microscopy (confocal excitation, two-photon excitation, near-field optics, fluorescence lifetime imaging), fluorescence correlation spectroscopy, and single-molecule fluorescence spectroscopy. 

Fluorescence spectroscopy and its applications to the physical and life sciences have evolved rapidly during the past decade. The increased interest in fluorescence appears to be due to advances in time resolution, methods of data analysis and improved instrumentation. With these advances, it is now practical to perform time-resolved measurements with enough resolution to compare the results with the structural and dynamic features of macromolecules, to probe the structures of proteins, membranes, and nucleic acids, and to acquire two-dimensional microscopic images of chemical or protein distributions in cell cultures. Advances in laser and detector technology have also resulted in renewed interest in fluorescence for clinical and analytical chemistry. 

Using flexible endoscopic catheters, intravascular fluorescence spectroscopy and imaging demonstrated that atherosclerotic plaques and other vascular abnormalities can be identified and characterized based either on endogenous fluorophores [11] or exogenous fluorescent dyes [146,147]. 

Combining fluorescence spectroscopy with fluorescence microscopy, confocal microscopy could be used to elucidate the pathway of siderophore-mediat iron uptake in the fungus Ustilago maydis, and visualize this pathway by providing unique fluorescent microscopic images. Using these techniques, clear images of two independent iron-uptake mechanisms have become visualized as well as their cellular compartment locahzed. 

Nevin A, Comelli D, Valentini G, Anglos D, Burnstock A, Gather S, Cubeddu R (2007) Time-resolved fluorescence spectroscopy and imaging of proteinaceous binders used in paintings. Anal Bioanal Chem 388 1897-1905. 

Figure 18-21 Excitation and emission spectra of anthracene have the same mirror image relation as the absorption and emission spectra in Figure 18-16. An excitation spectrum is nearly the same as an absorption spectrum. [C. M. Byron and T. C. Wemer. Experiments in Synchronous Fluorescence Spectroscopy lor the Undergraduate Instrumental Chemistry Course"...

Beside the identification of single bacterial cell or spores by means of Raman spectroscopy, the localization of these cells inside partly complex matrices has to be performed. One approach is the combination of Raman spectroscopy, fluorescence spectroscopy and digital imaging techniques. This method was applied to detect traces of endospores and other biothreat organisms even in the presence of complex environmental matrices like bioaeroso-lic background, nasal mucin [67], or tap water [68], Another fully automated device was built to analyze bioaerosols in clean room environments, where prior to the Raman identification method a particle preselection took place [69]. 

Basic principles and applications of time-resolved fluorescence spectroscopy have been outlined in a very illustrative way by Valeur [16]. Although punctiform spectroscopy is still the best way to get a detailed knowledge of all the important parameters that characterize fluorescence emission (exact spectral properties, decay time behavior, polarization), imaging is always preferred whenever the localization of the distribution of any biomolecule of interest is required or a great number of samples have to be analyzed [22]. 

Bastiaens PIH, Verveer PJ, Squire A, Wouters F (2001) Fluorescence lifetime imaging microscopy of signal transduction protein reactions in cells. In Valeur B, Brochon JC (eds) Springer series on fluorescence 1 (New trends in fluorescence spectroscopy). Springer, Berlin Heidelberg New York p 297-302... 

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