Modified fretting

Chip seal Loose chips from a freshly paved road are the major safety concern for chip seal operation, and several attempts were reported in the literature to develop a laboratory procedure to simulate the field experience. A modified fretting test (also know as the abrasion cohesion test Esso, ACTE) appears to be the most success-fid [26, 27]. In this test a known amount of CRS-2 asphalt emulsion and aggregates are spread on a roofing felt, and then rolled with a 301 rabber roller. The sample is subjected to the shearing action of a horizontal steam-hose, which is attached to a Hobart sun and planet mixer, and the percentage of retained chips is recorded as a function of curing time. 

Fig. 12-19 Results of modified fretting test demonstrating advantages of the early chip retention with cationic SBR latex modified emulsion.

Another important group of hydrolytic enzymes are phospho- and cyclophosphodiesterases. They catalyze the hydrolysis of phospho-diester bonds and many of the most relevant biological substrates are nucleic acids. Phospholipase C and D are also important examples. Initial attempts to measure phosphodiesterase activity placed a phosphodiester between a fluorophore and a quencher and the probe was tested in vitro [146], This system was slightly modified by Caturla and used for the identification of catalysts with phosphodiesterase activity [147], More recently, Nagano and co-workers used a coumarin donor and fluorescein as a FRET... 

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... 

Figure 10.3. Modified Jablonski diagram for the processes of absorption and fluorescence emission (left), dynamic quenching (middle), and fluorescence resonance energy transfer (FRET) (right).

Fig. 3. Kinetics of the different steps in GPCR activation and signaling. Shown are the fast and slow phases of PTH binding to the PTH receptor, and the PTH-induced increase in in tracellular cAMP. PTH binding was measured by FRET as indicated in Fig. 2A. cAMP levels were determined by FRET using the epacl-sensor (Nikolaev et at, 2004). Note that the conformational change of the PTH receptor (measured as depicted in Fig. 2B) has the same kinetics as the slow phase of PTH binding [modified from Castro et at. (2005)].

Although this convenient definition of (R)tw is somewhat different from that employed in the context of SM-FRET [Cf. Eq. (6)], one does not expect this distinction to modify the main observations reported below. 

We have developed the splinted RNA ligation procedure outlined in this chapter to generate site-specifically dye-labeled telomerase RNA constructs. These modified telomerase RNA constructs may be used to characterize dynamic RNA structural properties using Forster resonance energy transfer (FRET) (Stone et al., 2007). Our laboratory specializes in single molecule FRET measurements, which facilitates the direct observation of transient RNA structural states. The details of single molecule FRET... 

While the capture on DNA chips of fluorophore-labelled targets, and the extension of arrayed primers with fluorophore-labelled nucleotides has been widely used for some time, it is only more recently that assay formats have developed that utilize immobilized nucleic acids already modified with fluorophores. Fundamental analyses of surface monolayer structures and chemistries can be readily performed by immobilizing such modified oligonucleotides into SAM structures [105,106], but it is those interactions that can be monitored using fluorescence quenching or fluorescence resonance energy transfer (FRET) that have gained the most attention. 

Fig. 21 In situ visualization pictures showing damage a in a fretting contact between an unmodified DGEBA/DDM epoxy network and a glass sphere, and b in a similar contact with a DGEBA/DDM network modified with the acetamide derivative shown in Table 1. The magnitude of the maximum tensile stress at the edge of the contact was found to be similar in both cases. Contact fatigue cracks were only observed in the neat DGEBA/DDM system...

Chemically modified crowned spirobenzopyran 112, containing a pyrenyl fluorophore attached at the nitrogen atom, can function as a fluorescence emission switch <2004T6029>. This sensor displayed a quenching of the PET fluorescence emission of the fluorophore in the absence of metal ions (the merocyanine form was not produced). When, however, the spiro form of 112 was converted into the merocyanine form by metal ion complexation of the crown ether portion of the molecule, a fluorescence resonance energy transfer (FRET) from the pyrene to the merocyanine moiety took place, producing fluorescence emission. 

The FRET process is not the only factor that can modify the donor fluorescence lifetime. The local environment of the donor or its quenching by any compound of the library under investigation can also change its lifetime significantly and therefore disturb the FRET detection. 

There are also a number of new dye-modified analogues for use in FRET analysis. A new four colour set of FRET dideoxy 5 -triphosphate terminators has been developed involving a rigid, linear ET cassette linker chemistry, and formulated with Thermo Sequenase II DNA polymerase. An alternative approach for a four colour set of FRET dyes has been reported where the dyes are incorporated into ODNs as phosphoramidite derivatives. ... 

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