J. Hohlbein, M. Steinhart, C. Schiene-Fischer, A. Benda, M. Hof and C.G. Hübner, Small, 3, 380, 2007, [link]
Self-ordered nanoporous alumina was used to create a two-dimensional geometrical conﬁnement for either single diffusing molecules or ﬂuorescent polymerized nanowires. The membranes for measurements of single molecule diffusion featured a pore diameter of 35-40nm, a porosity (volume fraction of the pores) of 20-25%, and a thickness of 35 µm. Thus, the aspect ratio of the pores is ≃1000. In comparison, the size of the diffraction limited laser focus is roughly 2µm in height (long axis) and 0.5µm in diameter (short axes), resulting in an aspect ratio of 4. Therefore, if the long axis of the pores is aligned with the long axis of the confocal microscope, the probe molecules are forced to diffuse parallel to the long axis of the laser focus. Apparent one-dimensional diffusion within nanoporous alumina was shown for different probes such as Alexa Fluor 488 and the enhanced green ﬂuorescent protein (eGFP). As compared to three-dimensional diffusion in free solution, the mean diffusion time through the focus increases within the pores. The factor of increase was theoretically derived as the squared aspect ratio of the laser focus resulting in a value of 16. Indeed, for Alexa Fluor 488 an increase in the mean diffusion time by a factor of 19 was found. In the case of eGFP a factor of 14 was obtained.