Confocal Laser Scanning Microscopes have initiated a breakthrough in biomedical imaging. High contrast due to effective suppression of light scattered from outside the focal plane, simple fluorescence imaging by single photon or two-photon excitation and the 3D imaging capability are features beyond the reach of conventional microscopes. Adding ps time resolution by combining the confocal microscope with an advanced Time-Correlated Single Photon Counting (TCSPC) imaging technique yields a new powerful instrument for the investigation of molecular interactions in biological systems.

To investigate molecular interactions in cells and subcellular structures fluorescence markers are used which specifically link to protein structures. Staining the sample with different dyes and recording the fluorescence image reveals the cell structures via the different fluorescence spectra and fluorescence lifetime of the dyes. Energy transfer between the dye molecules and the proteins changes the fluorescence quantum efficiency and thus the fluorescence lifetime. Due to the variation of the dye concentration these effects cannot be distinguished in simple intensity images. Therefore, recording time-resolved patterns of the full fluorescence decay functions rather than simple intensity imaging is required to investigate molecular interactions in biological systems.

Recording time-resolved fluorescence images can be achieved by combining a confocal laser scanning microscope, a femtosecond Titanium Sapphire (TiSa) Laser and an advanced Time-Correlated Single Photon Counting (TCSPC) imaging technique.