FLIM Analysis Enhanced with Phasor Plotting aids Quantitative Biology

Fluorescence Lifetime Imaging (FLIM) is a powerful technique that provides a new dimension to quantitate the fluorescent probe, in addition to its steady-state intensity and spectral profiles. With particular selectivity of probes, FLIM can provide quantitative information about the probe microenvironment such as ions, pH, oxygen content, electrical signals, and the index of refractions.

FlIM is one of the most robust ways of quantifying F?rster Resonance Energy Transfer (FRET) for studying protein-protein interactions in live specimens. Turnkey commercial FLIM instruments are mature, making the technique easily accessible; however, a major challenge is still how to analyze and interpret FLIM data. The phasor plots approach has been demonstrated to be an effective solution. Here, several quantitative tools using the phasor plots in a multi-image phasor analysis (MiPA) module will be described; their utility will be demonstrated for various applications including time-resolved multiplexing, label-free NADH imaging, STED superresolution imaging, and FRET.

About the presenter
Yuansheng Sun, Ph.D., received his doctorate in biomedical engineering from the University of Iowa and was further trained in the Keck Center for Cellular Imaging at the University of Virginia. He is now working at ISS Inc. as director of microscopy and spectroscopy.