Frequency-domain fluorescence lifetime imaging (FD-FLIM).
Chong, Kau Sang.
Date of Issue2011
School of Chemical and Biomedical Engineering
As a non-intensity-based measurement technique, fluorescence lifetime imaging minimizes of photon scattering effect in thick layers of biological samples. Lifetime of different fluorophores can become the important source of characteristic information especially on the properties of the fluorescent molecules themselves or the changes in their microenvironment. Time-domain acquisition is commonly used in FLIM but it requires fast excitation pulse and detection circuit. Due to relatively rapid speed of data acquisition and simpler electronics, frequency-domain approach is applied in this research. The gain-modulated image intensifier coupled with CCD camera is the important optical instrument enabling the data acquisition in frequency domain. Despite the commonly-used absolute lifetime measurement, the experimental result shows high reliability in relative lifetime measurement, with two fluorescent dyes between which one is taken as reference to another, while the accuracy of lifetime from both demodulation,τm and phase shift,τφ are being investigated. We perform the experiment with free-space setup, from which a sensitive frequency-domain FLIM system is calibrated and prototyped, to gain the fundamental knowledge and to remove the complication that could arise from microscopy system.
Final Year Project (FYP)
Nanyang Technological University