KEYWORDS: Modulation, Phase shift keying, Luminescence, Monte Carlo methods, Cameras, Signal to noise ratio, Fluorescence lifetime imaging, Line scan image sensors, Imaging systems, Signal processing
We present a theoretical multi-frequency multi-sampling frequency-domain fluorescence lifetime imaging (FLI)
technique, which determines fluorescence lifetime by measuring the phase delay and decrease of the modulation of
the emission relative to the excitation. Such a system could be realized with high-speed line-scan cameras using
more than 100000 frames per second. It is useful for the measurement of lifetimes in the microsecond range and
can be, e.g., used to measure oxygen concentrations with Ruthenium complexes. Typically 100 samples are used,
resulting in a temporal resolution of microsecond fluorescence lifetime measurement. By including frequency
components with frequencies much higher than the Nyquist criterion, the measurement of fluorescence lifetimes
much shorter than the sampling interval is possible. In order to optimize the SNR of the predict lifetimes, we
investigate the photon economy of our technique. Various ranges of the exposure time relative to the range
of modulation period and compositions of multiple frequencies have been studied. The presented approach is
validated by numerical simulations by Monte Carlo method.
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