Fluorescence correlation spectroscopy in small cytosolic compartments depends critically on the diffusion model used

Fluorescence correlation spectroscopy (FCS) is a powerful technique for mea suring low concentrations of fluorescent molecules and their diffusion cons tants. In the standard case, fluorescence fluctuations are measured in an o pen detection volume defined by the confocal optics. However, if FCS measur ements are carried out in cellular processes that confine the detection vol ume, the standard FCS model leads to erroneous results. In this paper, we d erive a modified FCS model that takes into account the confinement of the d etection volume. Using this model, we have carried out the first FCS measur ements in dendrites of cultured neurons. We further derive, for the case of confined diffusion, the limits within which the standard two- and three-di mensional diffusion models give reliable results.