Fluorescence Lifetime Imaging (FLIM) and Time Correlated Single Photon Counting (TCSPC)

With the fluorescent lifetime imaging (FLIM) technique, the rate of decay of fluorescence is measured, rather than the intensity of fluorescent light. There are several methods of pulsed illumination that can be used to measure fluorescence decay, but the most common method is time correlated single photon counting (TCSPC).

This system is often used to look inside cells and tissues and has found particular use in the analysis of tumors and neurons. The key components of the FLIM with TCSPC system include a laser that excites the photon and a TCSPC unit that measures the rate of decay of that photon and records the measurement. This is done multiple times and the average results are calculated then reconstructed and displayed on a screen.

The benefits of FLIM include the fact that it is not concentration sensitive, the short measurement times, and a good signal-to-noise ratio, among other things. However, because light is used, the size of what can be imaged is diffraction limited. Fluorescence lifetime spectrometers are used to analyze organic compounds in medical and chemistry labs.