MICROSCOPE BASED FLUORESCENCE PHOTON-CORR ELATION

The contribution describes the development and the setup of a fluoresc ence correlation spectrometer (Zeiss ConfoCor(R)) based on an inverted light microscope (Zeiss Axiovert 135 TV(R)). The underlying measuring method is the fluorescence correlation spectroscopy (FCS). Starting w ith the theoretical and technical basis of the method, the requirement s which a FCS-system has to meet are explained. Emphasis will be laid on the confocal optical setup, on the single-photon detection system a nd on the correlation technique employed for the signal processing. Th e software FCS ACCESS(R) for the ConfoCor(R) runs under the operation system Microsoft Windows for Workgroups(R) or Microsoft Windows(R). Th e software package includes modules for the control of the instrument, for data processing and data presentation. For the analysis and evalu ation of the measured data non-linear optimization algorithms are util ized that result in a multi-parameter fit program. Analytical paramete rs as for example the number of molecules within the illumination volu me and the diffusion time are available within few seconds. From these parameters the binding constants and kinetic constants can be calcula ted. Two examples of measurements which are related to biomedical appl ications are described, namely the diffusion of rhodamine dye molecule s and the interactions of a receptor-ligand system.