Cyan fluorescent proteins CFPs

The analysis of the histograms of photon arrival times is equivalent in both cases and relies on fitting appropriate model functions to the measured decay. The selection of the fitting model depends on the investigated system and on practical considerations such as noise. For instance, when a cyan fluorescent protein (CFP) is used, a multi-exponential decay is expected furthermore, when CFP is used in FRET experiments more components should be considered for molecules exhibiting FRET. Several thousands of photons per pixel would be required to separate just two unknown fluorescent... 

Cyan fluorescent proteins (CFPs) have blue-shifted excitation and emission spectra, because of the mutation Tyr66Trp inside the chromophore (Fig. 5.3C) [34], CFP fluorescence (Ex 435 nm/Em 474 nm) is less blue-shifted than for EBFP and CFP excitation is intermediate to the excitation of the neutral and anionic chromo-phores of avGFP [4], CFPs are widely used for dual-color imaging and FRET applications together with yellow fluorescent proteins (YFP, Section 3.6). 

FRET requires the presence of two fluorophores, one with a shorter emission wavelength (donor) and another with a longer emission wavelength (acceptor). The fluorophores must be chosen such that there is sufficient overlap of the donor emission spectrum and the acceptor excitation spectrum. When FRET occurs, which requires the proximity of the two fluorophores, excitation of the donor results in transfer of energy to the acceptor and, hence, emission at the wavelength characteristic for the acceptor. FRET can be seen with various kinds of fluorophores, but most recendy it has been used in particular with variants of GFPs because this permits FRET in intact cells. The most frequently used pairs of GFPs are the cyan fluorescent protein (CFP) and the yellow fluorescent protein (YFP) variants. The donor CFP is excited at its maximum... 

The first described member of the cyan fluorescent proteins (CFPs) resulted from a rationally designed chromophore mutation of Aequorea GFP. Heim and co-worker replaced Tyr66 with Trp and found the peak wavelength for excitation and emission of this GFP derivative (GFP-Y66W) to be shifted to 436 and 476 nm, respectively [52], Because of this blue-green/cyan light emission the protein was called cyan fluorescent protein or CFP. 

Table 4. GFP-derived cyan fluorescent proteins (CFPs)...

CFP Cyan fluorescent protein. CFP is a derivative of the green fluores-... 

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