Experimental Setup

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The “classical approach” for FRAP measurements utilizes a widefield epi-fluorescence microscope to monitor the fluorescence intensity during pre- and postbleach acquisition. For bleaching a laser beam is focused onto a small diffraction limited spot (~1µm) in the region of interest. These systems are not ‘of the shelf’ and have to be custom-build.

In principal the standard commercially available confocal laser-scanning microscope (CLSM) equipped with acousto-optical tuneable filter (AOTF) is suitable for FRAP, iFRAP and FLIP experiments. The AOTF is needed to switch the laser power rapidly between the low intensity imaging and the high intensity bleaching modes.

The higher the diffusion constant of the investigated molecule, the more imaging speed and laser power is required. Read more about useful confocal equipment for FRAP experiments.

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contact: Stefan Terjung           last update: 02/06/04