Fine resolution. By using an aperture with a subwavelength diameter at the end of the probe, the diffraction limit of conventional light microscopy ( / 2) is avoided, and a finer-resolution structure can be visualized.

Lipofuscin accumulates as yellow-brown granules in the retinal pigment epithelium (RPE) of the human eye and can take up significant cell volume in older individuals. It has been proposed that the substance plays a role in the development of age-related macular degeneration, a major cause of blindness. Lipofuscin is believed to be a photochemical source of reactive oxygen species that form under blue light and lead to RPE damage.

The international team of researchers is determining the distribution of the orange fluorescent chromophores on the surface of the granules to assess their importance to the oxygen activation process. To accomplish this, they applied a recently developed technique called near-field scanning optical microscopy (NSOM) to lipofuscin granules collected from older people. They funneled light along the lipofuscin surface from about 10 nm away with an optical fiber that is smaller in width than the wavelength of the released light. Initially, expelled light contains highly resolved waves that are smaller than the specified wavelength of the light. These waves dissipate almost immediately, but the distance between the fiber and the surface is short enough to allow them to interact with the sample. This process results in very well resolved topographical and emission images of the lipofuscin granules.

The scientists determined, with high resolution, the fluorescence distribution over the surface structure of several individual granules. They found that the orange fluorophores are not homogeneously distributed. The implications of their specific measurements are under investigation. The team plans to perform photochemical oxygen activation experiments on spatially selected regions of the lipofuscin granules using NSOM.