Intensity fluctuations, fluorescence

The dye-loaded dendrimer 110 can be detected by means of single-molecule fluorescence.48,57,281,282 This multichromophoric system was embedded into a thin polymeric film at very low concentrations so that individual dendrimer molecules could be studied by far-field fluorescence microscopy at room temperature. Temporal fluorescence intensity fluctuations, fluorescence spectra, and decay times were measured separately or simultaneously. Collective on/off jumps of the fluorescence intensity as well as different emissive levels were observed for single dendrimer molecules with eight perylenemonoimide chromo-... 

Common to most fluorescence-based single-molecule methods, photobleaching limits the observation time window using the fluorescence quenching via energy transfer approach. In addition, this approach only obtains the fluorescence intensity from one probe thus, fluorescence intensity fluctuations due to probe photophysics, such as fluorescence blinking, can complicate the results and data analyses. Triplet quenchers such as Trolox 95 can reduce fluorescence blinking. Careful control experiments are in any case necessary. 

The quantity measured in FCS experiments is the autocorrelation function of the fluorescence intensity fluctuations, G(t), defined as ... 

A third method for measuring spectral dynamics of individual molecules in glasses involves fluorescence intensity fluctuations during steady-state excitation at a fixed frequency. [20] This method has been applied to the systems of Tr in PE [18,20] and TBT in PIB [15, 16]. In these experiments the fluorescence intensity fluctuates as the chromophore moves in and out of resonance because of coupling to flipping TLSs. Thus this very clever technique can provide a direct probe of TLS dynamics on all time scales. 

The third type of experimental observable for single molecule spectral dynamics is the correlation of fluorescence intensity fluctuations. In this experiment one pumps the chromophore under steady-state conditions at a fixed frequency g>l and measures the time-dependent intensity /(l), whose fluctuations are analyzed by the correlation function... 

Extremely exciting experimental data for glasses are now beginning to emerge. It has been shown that line shape measurements, fluorescence intensity fluctuations, and spectral diffusion trajectories can all be used to probe TLS dynamics on different time scales. Furthermore, as has been emphasized already, these experiments on individual molecules will provide information complementary to that obtained from more traditional echo and hole burning experiments. At this point what we need is more data. In an ideal world all three of the above experiments would be performed on the same individual molecule at a variety of temperatures, and then would be repeated on many molecules, and all of the above would be repeated for several different systems. Although the basic theoretical apparatus is in place for analyzing these experiments, more refined theoretical results will surely be needed. 

Ehrenherg M, Rigler R (1974) Rotational Brownian-motion and fluorescence intensity fluctuations. Chem Phys 4(3) 390 01... 

Intramolecular Dynamics of Chain Fluorescence Intensity Fluctuations... 

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