Fluorescence intensity regime

It is possible to also write expressions for the modified quantum yield and fluorescence intensity enhancement factor in the UFDMEF regime. These are similar to those obtained for the SDMEF regime. 

In this method, two pulsed lasers are used, both usually in the nanosecond regime. One (the burn laser) is operated at high power, and is scanned across the absorption spectrum. It excites molecules (or clusters) from the particular vibrational level (usually the i = 0 level) to an electronically excited state. The upper state relaxes (radiatively or otherwise) back to the ground state, but not necessarily to i = 0. Thus, depletion in the population of this species is achieved. A second, low-power laser (the probe laser) is fired after a suitable time delay (to allow complete decay of the emission induced by the pump laser). It is tuned to one of the excitation spectrum vibronic bands of the system, and the fluorescence induced by it (the signal ) is continuously monitored. Whenever the frequency of the bum laser corresponds to excitation of the species giving rise to the absorption of the probe laser, the signal is reduced. This reduction appears as a hole that is burned in the spectrum—hence the name of the method. If a different species is excited (another molecule or a different vibrational level) no change in fluorescence intensity is incurred. 

If now the period T of the pulses changes and becomes smaller or greater than the atomic period T] 2> it produces dephasing between the various atomic responses S (t) and blurring of their summation the total fluorescence intensity, in stationary regime, is modulated with a smaller amplitude. The amplitude of the fluorescence modulation passes through a maximum when the excitation frequency a> becomes equal to the atomic frequency 12 practice, it is not necessary to use a pulse excitation it is necessary to use a broad band excitation (Aa)exc 12 and to interrupt periodically the excitation light 13. ... 

The incoherent transfer for t > 10 ps has first a multi--exponential character which becomes at t 25 ps a single--exponential one. The single-exponential transfer for t > 25 ps is the most important one as it gives the most significant contribution to 7)0j (about 90 %). In this regime, the fluorescence intensity and other observables have single--exponential character. The corresponding life time equals... 

In order to understand the influence of different molecular parameters on structure formation and stability, we report on the directed encapsulation of two hydrophobic model substrates inside the polymersome shell. The system both complies with requirements like water insolubility and has sufficient fluorescence intensity for monitoring. Fluorescent CdSe/CdS/ZnS core-shell quantum dots (QDs), which carry hydrophobic surface ligands, serve as a model substrate of the nanosize regime (core size approximately 6 nm). Nile Red, a lipophilic fluorescent dye, represents the molecular size regime [268-270]. 

In a recent paper by Vigud and co-workers [7] coherent saturation effects in laser-induced-fluorescence are discussed. Coherent effects should be considered in situations where the laser coherence time is comparable to the time the molecule interacts with the laserfield. In the saturation regime where the Rabi ffequency co/ is much larger than the total decay rate out of the excited state the fluorescence intensity turns out to be proportional to (O/j, i.e. proportional to the square root of the laser power. 

We sh( uld point out that these experiments were done in a totally different (XHicentration range compared to the hybridization studies for the LOD determination (me ( lerates in the mass transfer-limited regime of the binding process. I.e., the concentrati(m of the analyte is so low that one observes only the diffusion-controlled approach of labeled analyte molecules probed by the SP field giving rise to a linear increase of the fluorescence intensity with time. 

The fluorescent components are denoted by I (intensity) followed by a capitalized subscript (D, A or s, for respectively Donors, Acceptors, or Donor/ Acceptor FRET pairs) to indicate the particular population of molecules responsible for emission of/and a lower-case superscript (d or, s) that indicates the detection channel (or filter cube). For example, / denotes the intensity of the donors as detected in the donor channel and reads as Intensity of donors in the donor channel, etc. Similarly, properties of molecules (number of molecules, N quantum yield, Q) are specified with capitalized subscript and properties of channels (laser intensity, gain, g) are specified with lowercase superscript. Factors that depend on both molecular species and on detection channel (excitation efficiency, s fraction of the emission spectrum detected in a channel, F) are indexed with both. Note that for all factorized symbols it is assumed that we work in the linear (excitation-fluorescence) regime with negligible donor or acceptor saturation or triplet states. In case such conditions are not met, the FRET estimation will not be correct. See Chap. 12 (FRET calculator) for more details. 

The case of MB in the SDMEF regime (Fig. 2.2(b)) is typical of many MEF experiments. Moreover, to maximize the MEF intensity, the LSP resonance is typically chosen in between the laser excitation and the peak fi-ee-space fluorescence emission, i.e. cases B and C in Fig. 2.2(b). Only small SPMs are predicted in these conditions a small shift for B and a small narrowing for C. These situations are in fact representative of typical MEF conditions and small SPMs should therefore (in general) be expected. Such small modifications have indeed been reported [11], but it is likely that they remain unnoticed or have not been emphasized as such in many cases. 

---