Fluorescence decay data analysis correcting for detector pulse pile-up at very high count rates
arXiv:1711.01137 · doi:10.1117/1.OE.57.3.031305
Abstract
Using Time-Correlated Single Photon Counting (TCSPC) for the purpose of fluorescence lifetime measurements is usually limited in speed due to pile-up. With modern instrumentation this limitation can be lifted significantly but some artefacts due to frequent merging of closely spaced detector pulses (detector pulse pile-up) remains an issue to be addressed. We propose here a data analysis method correcting for this type of artefact and the resulting systematic errors. It physically models the photon losses due to detector pulse pile-up and incorporates the loss in the decay fit model employed to obtain fluorescence lifetimes and relative amplitudes of the decay components. Comparison of results with and without this correction show a significant reduction of systematic errors at count rates approaching the excitation rate. This allows quantitatively accurate fluorescense lifetime imaging (FLIM) at very high frame rates.
12 pages, 3 figures. v2: spelled out unexplained abbreviation FRET as requested by reviewer. v3: updated acknowledgements, accepted for publication in Opt. Eng