Intelligent Imaging Innovations (3i) FLIM system is easy to use and can be combined with many different microscopy applications, such as widefield, deconvolution, spinning disk confocal, and TIRF. It is based on wide field frequency-domain homodyne detection, which uses a high frequency modulated laser light source and synchronized intensifier. A lifetime image can be acquired in less than a second, making it significantly faster than a point scanning photon counting system which is particularly useful for monitoring FLIM in living cells. FLIM analysis is closely integrated into 3is SlideBook software. Through an intuitive polar plot (phasor) graphical analysis, it is easy to visualize multiple fluorescent species, FRET efficiency, and mixture analysis. Numerical tools complement the visual analysis, and provide first- and second-order FRET measurement, nearest single-exponential maps, and graphs of lifetime (or FRET) versus time which can be displayed during capture. SlideBook analysis can be further customized through its interface to MATLAB.