We evaluated fluorescence microscopy combined with digitized image analysis for the investigation of fluorescein transport in the intact rat liver. The images of the surface of isolated rat livers which were perfused directly under a microscope were projected onto a silicone-intensified target camera, stored on a video tape and analyzed with a microcomputer equipped with an image digitizer. It was shown that, after correction of the day-to-day variability of the optical and electronical system, the increase in fluorescence intensity of the liver surface following fluorescein infusion depended linearly on the fluorescein concentration in the perfusion medium up to 1 mM. Since, under the conditions used, biliary secretion and metabolic influences were found to be insignificant the uptake mechanism is probably predominantly simple diffusion.