Signals obtained in fluorescence lifetime imaging microscopy (FLIM) often contain biexponential decay components, and unmixing of them is a critical issue in FLIM especially for fluorescence resonance-energy-transfer (FRET) applications. Even though the analog mean-delay (AMD) method is the fastest fluorescence lifetime measurement scheme for point-scanning confocal microscopy, its ability to analyze biexponential decay signals has not been demonstrated, yet. In this paper we have demonstrated that biexponential decay signals can be effectively analyzed with the AMD method. Effects of various measurement conditions on the accuracy of biexponential decay signal analysis are presented with numerical simulations and experimental demonstrations.
All Science Journal Classification (ASJC) codes
- Electronic, Optical and Magnetic Materials
- Atomic and Molecular Physics, and Optics
- Physical and Theoretical Chemistry
- Electrical and Electronic Engineering