Method of Simultaneous Frequency-Sweeping Lifetime Measurements on Multiple Excitation Wavelengths

摘要

A fast fluorescence lifetime microscopic system images FRET between multiple labels in live cells and deep tissue, using a quantitative analysis method to reconstruct the molecular machinery behind the multiplexed FRET phenomenon. The system measures fluorescence lifetime, intensity and anisotropy as images of excitation-emission matrices (EEM) in real time and high speed within a single image scan, performs high-resolution deep-penetrating 3D FRET imaging in live samples, and fully analyzes all possible photon pathways of multiplexed FRET. The system provides a way for systematic and dynamic imaging of biochemical networks in cells, tissue and live animals, which will help to understand mechanisms of genetic disorders, cancers, and more.