Confocal laser scan fluorescence

McConnell, G. (2004). Confocal laser scanning fluorescence microscopy with a visible continuum source. Opt. Express 12, 2844—50. 

Confocal laser scanning fluorescence microscopy was used to study the exposure of the avidin-specific binding sites in the Av-GEB platform by the immobilization of a small and flexible biotinylated fluorescein molecule as a fluorescence marker. Fluorescence microscopy thus confirms that Av-GEB platform exposes active binding sites for biotin, acting as affinity matrix (Fig. 21.2B). After use, the electrode surface can be renewed by a simple polishing procedure for further uses, highlighting a clear advantage of this new material with respect to surface-modified approaches such as classical biosensors and other common... 

Fig. 21.2. (A) Schematic representation of the electrochemical DNA biosensing procedures based on Av-GEB. (B) Confocal laser scanning fluorescence microphotograph of Av-GEB transducers submitted to (i) non-biotinylated fluorescein (background adsorption) and (ii) 80 pmol of biotinylated fluorescein. Laser excitation was at 568 nm. Voltage 352 V (more details in Zacco et at., [65]).

For biotechnological purposes, it is necessary not only that cells remain viable but also that adhesion is maintained so that a cell can be found in the expected position. The adhesion pattern that a cell has adopted before freezing is crucial to successful cryo-preservation. Adhesion patterns can be visualised by (TIRF microscopy, [63]) mentioned above and also by confocal laser scanning fluorescence (CLSM, [64]) microscopy. We have investigated the adhesion patterns of fibroblasts as a function of adhesion time and various surface treatments [65]. Some examples are shown in Fig. 19. 

Other methods of imaging fat crystals and fat crystal networks (not all of them optical) include confocal laser scanning fluorescence microscopy, multiple photon microscopy, atomic force microscopy and electron microscopy (Narine and Marangoni, 1999). 

In recent years, many other techniques have been employed to elucidate the structure of fat crystal networks including confocal laser scanning fluorescence microscopy (Heertje et al. 1987) and multiple photon microscopy (Marangoni and Hartel 1996). Another advance has been the development of three-dimensional imaging. 

The last two decades have seen tremendous progress in optical microscopy, especially, confocal fluorescence microscopy. Confocal laser-scanning fluorescence... 

Confocal laser-scanning fluorescence microscopy (CLSM). The micrographs were obtained by means of a Leica confocal scanning system mounted to a Leica Aristoplan. A 1 OOX oil immersion objective with the numerical aperture 1.4 was used. The standard filter settings for fluorescent excitation and emission were used. 

Figure 1.1 Confocal laser scan fluorescence images of single crystals of (a, b)...

Guthoff RF, Wienss H, Hahnel C et al (2005) Epithelial innervation of human cornea. A three-dimensional study using confocal laser scanning fluorescence microscopy. Cornea 24 608-13... 

The latter process was performed by immersing the patterned amino polymer films into a solution containing FITC, then the film was intensively washed, and the resulting fluorescent structure finally was visualized with a confocal laser scanning fluorescence microscopy. The result is displayed in figure 3b). 

With the advent of confocal laser scanning fluorescence microscopy (CLSFM) [39], multiple photon microscopy (MPM) [40,41], and atomic force microscopy (AFM) [5], three new tools have been added to the standard tools of light microscopy (LM) [42-45] and electron microscopy (EM) [35,46-50] that were most widely used in the past to study the microstructure of fats and foods in general. The work of Heertje et al. [35,36,39,51] on visualization of the microstructure in fats remains one of the most important contributions to the field. In their method, a cold solvent mixture (butanol-methanol) was used to remove the liquid oil form the solid fat in a sample mounted on a special holder. After removal of the liquid oil, the structure of the solid fat network could be visualized. 

Fig. 10 Confocal laser scanning microscope images of islets with urokinase (UK) immobilized on the membrane. The green fluorescence indicates positive immunostaining for UK. (a) Islets were modified with oligo(dT)2o-PEG-lipid (C16) or (b) oligo(dT)2o-PEG-lipid (C18) then, oligo (dA)2o-UK was added to the media, (c) Unmodified islets with (left) and without (right) oligo (dT)20-PEG-lipids added to the solution. Insets. Bright field images. Scale bars 100 pm...

Quader H. Formation and disintegration of cistemae of the endoplasmic reticulum visuahsed in hve cells by conventional fluorescence and confocal laser scanning microscopy evidence for the involvement of Ca2+ and the cytoskeleton. Protoplasma 1990 155 166-175. 

Two-photon excitation provides intrinsic 3-D resolution in laser scanning fluorescence microscopy. The 3-D sectioning effect is comparable to that of confocal microscopy, but it offers two advantages with respect to the latter because the illumination is concentrated in both time and space, there is no out-of-focus photo-bleaching, and the excitation beam is not attenuated by out-of-focus absorption, which results in increased penetration depth of the excitation light. 

Fig. 7.6. A and B. Confocal laser scanning microscographs of schistosome adults showing auto-fluorescence 488 nm was used for excitation and 520 nm for emission. A. Mid-body of a male schistosome with the focus on the lumen of the bipartite gut (g). In bombarded worms, a pale auto-fluorescence sometimes occurs under in vitro culture conditions. This auto-fluorescence occurs over a wide spectrum of wavelengths.

Confocal laser scanning microscopy can be used in conjunction with microwave heating for examining the three-dimensional structure and cellular interrelationships in sections of paraffin-embedded tissues (Boon and Kok, 1994). Tissues are fixed with Kryofix, a coagulant fixative containing 50% ethyl alcohol and polyethylene glycol (PEG molecular weight 300) for 90 sec in a microwave oven. The use of thick paraffin sections (15 (xm) and fluorescently labeled antibodies is preferred. 

Meeuwissen, M.E.M.J., et al. 1998. A cross-section device to improve visualization of fluorescent probe penetration into the skin by confocal laser scanning microscopy. Pharm Res 15 352. 

The distribution of LCM in the brain parenchyma was further analyzed using fluorescently labeled LCM and confocal laser scanning microscopy. As in the case of unlabeled LCM, rats bearing 9L gliosarcoma tumors were injected intravenously (i.e., via tail vein) with diO-LCM and sacrificed 2 min later. The brains were processed as described elsewhere (ref. 531). In this case,... 

Fig. 13.3. This figure demonstrates the distribution of fluorescently tagged LCM in brain parenchyma analyzed by confocal laser scanning microscopy. Rats bearing 9L tumors were administered diO-LCM and sacrificed 2 minutes later. Vibratome sections were counterstained with TR-WGA which binds to tumor cells and distinguishes the tumor area from the surrounding normal tissue. Comparison of the area stained with TR-WGA (tumor cells) (B) and that stained with diO (A) indicates that LCM were associated with a large portion of the tumor. (Taken from ref. 531.)...

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