This presentation introduces : The principles of Phasor analysis of Fluorescence Lifetime Microscopy (FLIM) data The use of FLIM of endogenous biomarkers for metabolic imging and the open source software to analyze FLIM data: FLUTE – (F)luorescence (L)ifetime (U)ltima(T)e (E)xplorer: a Python GUI for interactive phasor analysis of FLIM data The software is available on GitHub: https://github.com/LaboratoryOpticsBiosciences/FLUTE and it is published on Biological imaging Journal: Gottlieb, D., Asadipour, B., Kostina, P., Ung, T., & Stringari, C. (2023). FLUTE: A Python GUI for interactive phasor analysis of FLIM data. Biological Imaging, 1-22. doi:10.1017/S2633903X23000211 The lecture was part of the MIFOBIO 2025 Workshop A-07 Analyzing FLIM data for metabolic imaging applications with Phasor Analysis