Phasor theory of fluorescence lifetime imaging utilized on a maximum range of frequencies for prostate tissue analysis

Fluorescence lifetime imaging (FLIm) can detect macroscopic tumor tissue in various organs by measuring tissue autofluorescence, making it a compelling tool for surgical guidance. However, the fluorescence lifetime characteristics of tissue autofluorescence are complex due to the unpredictable microenvironment of the biomolecules in tissue, which complicates data interpretation. Nevertheless, the phasor analysis method is computationally fast and easily interpretable, making it appealing for clinical applications of FLIm. While many implementations of the phasor analysis operate only at a single frequency or a few harmonic frequencies, the phasor theory applied to pulse sampling FLIm as presented in this study leverages the maximum amount of frequency information, thereby extending the set of features available for tissue characterization. The clinical effectiveness of utilizing the maximum range of frequencies in phasor theory applied to pulse-sampling FLIm is demonstrated by investigating tumor detection in ex vivo tissue from 12 patients with prostate cancer. By accounting for the zonal anatomy of the prostate, it is shown that the degree of separability between healthy and tumor tissue is a function of frequency, and hence, the ability to access arbitrary frequency content can improve tumor detection in clinical guidance.