Dye Texas Red

Fig. IV-19. Fluorescence micrographs showing the shape transitions in monolayers of dimyristoylphosphatidylcholine (DMPC) (84%) and dihydrocholesterol (15%) and a lipid containing the dye, Texas Red. (From Ref. 228.)...

Since endocytosis ofLDH was confirmed by TEM images (Figure 13.9), forthe next step, its specific endocytic pathway for membrane entry was determined by immunofluorescence and confocal microscopy. Cells were incubated with LDH-FITC, fixed with 3.7% freshly made formaldehyde, and then stained with either anti-clathrin antibody or anti-caveolin-1 antibody both conjugated to the red fluorescent dye Texas Red (TR). The confocal microscopic images showed that green fluorescent... 

Alternatively, proteins can be labeled selectively using amine-reactive dyes. Particularly, cysteine and lysines can be modified covalently with a variety of commercially available fluorophores including Texas Red, Oregon Green, and Cy3 [19] (see also Chapter 6 for small molecule FRET probes, and Chapter 12 depicting a variety... 

In addition to the wide range of commercial probes, many other fluorescent molecules have been synthesized and described in the literature. Only a handful, however, are generally used to label antibody molecules. Perhaps the most common fluorescent tags with application to immunoglobulin assays are reflected in the main derivatives produced by the prominent antibody manufacturing companies. These include derivatives of cyanine dyes, fluorescein, rhod-amine, Texas red, aminomethylcoumarin (AMCA), and phycoerythrin. Figure 20.16 shows the reaction of fluorescein isothiocyanate (FITC), one of the most common fluorescent probes, with an antibody molecule. 

Albert H. Coons was the first to attach a fluorescent dye (fluorescein isocyanate) to an antibody and to use this antibody to localize its respective antigen in a tissue section. Fluorescein, one of the most popular fluorochromes ever designed, has enjoyed extensive application in immunofluorescence labeling. For many years, classical fluorescent probes such as FITC or Texas red (TR) have been successfully utilized in fluorescence microscopy. In recent decades, brighter and more stable fluorochromes have continually been developed (see Table 14.1). Marketed by a number of distributors, cyanine dyes, Cy2, Cy3, Cy5, Cy7, feature enhanced water solubility and photostability as well as a higher fluorescence emission intensity as compared to many of the traditional dyes, such as FITC or TR. 

Fig. 16 Addition of 0.017-nmol aliquots of a rhodamine B-labeled streptavidin and b Texas Red-X-labeled streptavidin to 1.51 nmol of 43. Energy transfer observed in both cases with amplified emission of the dyes to the light-harvesting conjugated polymers. Direct excitation of the dyes at 575 and 585 nm correspond to 0.100 nmol of streptavidin.

The interactions of the dye acceptors to polymers could also be witnessed in the case of solid films. In this case, the sterically more restrictive cavities of the polymeric film 46 allowed better orbital interaction with the smaller and more flexible rhodamine B dye, and accordingly higher energy transfer with rhodamine B-labeled streptavidin was observed compared to Texas Red-... 

The potential for multi-fluorochrome analysis has increased further with the availability of dyes excited by shorter wavelengths (such as Cascade blue) and longer wavelengths (such as Allophycocyamn and Texas red). With multiple laser flow cytometers, five cell-surface antigens (1) or more may be analyzed simultaneously. In principle, these methods are a natural extension of the methods outlined here, but require more sophisticated flow cytometers, and much more powerful computer facilities to explore the exponentially more complex data sets arising from such multiparameter analyses. 

Furthermore, some of the older, nonbranded, dyes, for example, Texas Red (Ha et al, 1996), fluorescein (Xie et al, 2004), or tetramethyl rhoda-mine (Lang et al, 2004), can work perfectly well in single molecule applications, and these dyes typically cost less than those noted above. 

Additional studies were performed where antisense ODN to the 6-opioid receptor was conjugated to Texas Red dye in order to visualize neuronal... 

Figure 2. Cultured pulmonary artery endothelial cells stained for tubulin (red), actin (green) and DNA (blue). The dual immunofluorescence procedure used rabbit anti-actin IgG and mouse anti-alpha tubulin IgG as primary antibodies. The secondary antibodies used were Texas Red-conjugated goat, anti-rabbit IgG and FITC-conjugated goat, anti-mouse IgG. The sample was also stained with the DNA-specific dye Hoechst 33342. Scale bar is equal to 20 microns.

N-acetyltyrosinamide (green), and N-acetylphenylalaninamide (red) in aqueous solution. The absorption spectra (left) show the relative absorption strengths of the three amino adds. The fluorescence spectra (right) are normalized to the same peak intensity (see Table 1 for relative quantum yields), (b) Fluorescence emission spectra of the malelmide derivatives of four common fluorescent dyes Alexa Fluor 488 (green), tetramethyirhodamlne (yellow), Texas red (orange), and Cy 5 (red). 

Duochrome has yet to find widespread use, and much of the work with three antibodies is undertaken using two lasers (2,3). An argon-ion laser is used to excite fluorescein and PE, whereas a second laser is used to excite the third dye. Either Texas Red is excited at 568 nm by a krypton or a dye laser, or allophycocyanin, another phycobiloprotein, is excited by a helium-neon laser at 630 nm. 

By attaching a donor-acceptor TMR/Texas Red pair of dye molecules site-specifically to noninterfering sites on the enzyme (Fig. 24.1a) [12], we were able to measure the hinge-bending motions of the enzyme by monitoring the donor-acceptor emission intensities (Fig. 24.1b) as a function of time. We estimated the Forster distance Rq [19] of a TMR/Texas Red pair to be about... 

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