Advancements in phasor-based FLIM: multi-component analysis and lifetime probes in biological imaging

Area of Science:

• * Biophysics
• * Optical Microscopy
• * Spectroscopic Imaging

Background:

• * Fluorescence lifetime imaging microscopy (FLIM) provides temporal data complementing intensity measurements.
• * Advancements in FLIM technology have expanded its applications.
• * Efficient processing of FLIM data is key to improving imaging speed and accuracy.

Purpose of the Study:

• * To review lifetime probes used in phasor-based FLIM.
• * To discuss the role of these probes in phasor analysis (PA).
• * To explore multi-component PA methods in biological settings.

Main Methods:

• * Phasor analysis (PA) for direct, fitting-free processing of lifetime data.
• * Utilization of lifetime probes with distinct lifetimes for visualization and cluster analysis.
• * Application of phasor vector rules and unmixing for multi-component analysis.

Main Results:

• * PA offers superior speed compared to traditional fitting algorithms for FLIM data.
• * Lifetime probes combined with PA facilitate visualization and cluster analysis.
• * Multi-component PA methods enable analysis within complex biological environments.

Conclusions:

• * Phasor-based FLIM, utilizing specific lifetime probes, provides rapid and intuitive detection.
• * This approach enhances the analysis of complex biological systems.
• * PA integration significantly improves the speed and visualization capabilities of FLIM.