Calculating FRET lines

A FRET line is a projection of a parameterized model to a set of single-molecule FRET observables. Such FRET experiment typically jointly determined the fluorescence weighted average lifetime of the donor \(\langle \tau \rangle_f\) in the presence of an acceptor and the FRET efficiency, E, for every molecule. Next, a multidimensional frequency histogram of these […]

Time-resolved detection: Beauty and benefits

Time-resolved fluorescence spectroscopy experiments track the emission of fluorescence light from a sample. Time-resolved fluorescence intensities are also called fluorescence decays. Analysis thereof may resolve composite samples and report on dynamics (citations). Excited fluorescent molecules stay a particular time in their excited state before emitting light and delay the detection of fluorescence. This “delay” depends […]

Fluorescence decays: Art and artifacts

One might think the analysis of a fluorescence decay is easy. In the end, it’s just standard curve analysis. However, the art is to precisely describe the fluorescence decay down to the shot-noise of the experiment. When doing so, a number experimental aspects have to be considered for the analysis: the instrumental response function, IRF, […]