Imaging techniques using fluorescence as the source of contrast has been widely
used in the last decades in biology. Even though fluorescence can give a lot of
information, it is sometimes hard to see contrast based on the intensity of
fluorescent molecules. This bachelor thesis is based on the idea that contrast in
fluorescence images can also be obtained using something different than intensity.
In this project, the source of contrast will be the lifetime of a fluorophore, which is
understood as the time it takes for an excited electron to go back to its ground state.
This property is not affected by concentration, photobleaching or quenching.
However, it can vary depending on the microenvironment pH, temperature,
viscosity, or binding to other molecules. All of these characteristics make lifetime a
very suitable property for studying different dynamic processes in the cells, at the
same time as being a good source of contrast in highly autofluorescence samples.
The main objective of this thesis was to build a system based on frequency domain
lifetime imaging. For its development intensity modulation of a laser as a sine wave
at MHz frequencies was required, as well as precise coordination between the laser
and the acquisition system (ICCD camera) to see contrast based on lifetime. The
project was performed at Universidad Carlos III de Madrid where the
implementation of the system took place, as well as its testing to check its capacities
and limitations.Ingeniería Biomédic
