Slides of the PoL-Bio-Image Analysis Symposium 2023: napari-flim-phasor-plotter: a Plugin to Generate Interactive Phasor Plots from Raw FLIM Data

<p>Fluorescence Lifetime Imaging Microscopy (FLIM) is a powerful technique used in various research<br>fields to study dynamic processes within biological samples. Among FLIM methods, Time-Correlated<br>Single Photon Counting (TCSPC) is widely employed due to its high sensitivity and accuracy in<br>measuring fluorescence lifetimes. TCSPC captures photon arrival times, allowing precise lifetime<br>determination. Researchers utilize FLIM to investigate molecular quantities like metabolism, FRET, pH,<br>and ion concentrations, offering new contrast in images. However, analyzing complex fluorescence<br>decays conventionally requires fitting models, which can be challenging. The Phasor Plot approach<br>revolutionizes FLIM data analysis by transforming complex fluorescence decays into a graphical<br>representation. The Phasor Plot is a polar plot, where each pixel's fluorescence decay is represented<br>by two coordinates. The plot provides a straightforward visualization of lifetimes and enables<br>unmixing different lifetime components without the need for fitting, making it a valuable tool for biological studies. A few different software were developed capable of analyzing FLIM data over the<br>last decades, however reading the raw FLIM data remains a non-trivial task for non-programmers.<br>We developed an open-source napari plugin called napari-flim-phasor-plotter, available<br>here: https://github.com/zoccoler/napari-flim-phasor-plotter. It is capable of opening and allowing<br>visualization of raw FLIM TCSPC data and generating the Phasor Plot inside the napari viewer. Since<br>the raw data can be very large, it can convert them to .zarr file format to allow lazy loading and<br>computing using the dask library. Beyond that, it integrates with another popular napari plugin, the<br>napari-clusters-plotter, thus, it opens the door for clustering algorithms to step into the universal<br>semi-circle. Segmentation results can be directly obtained from the phasor plot either manually, or<br>by means of a clustering algorithm.</p>