Achievement

IGERT participants at the University of Colorado and collaborators at Stanford University demonstrated a method for 3D optical imaging of objects smaller than 20 nm over a wide spatial range-overcoming the optical diffraction limit by an order of magnitude. The approach uses single fluorescent molecules and a wide-field microscope. The researchers engineered the microscope’s point-spread function (PSF) to have two points of light in the image plane. Because the orientation of the spots changes depending on the axial position of the molecule, the PSF appears as a double-helix along the axis of the microscope. In the experiments, single fluorescent molecules were localized within 20 nm over a large depth of field in a thick polymer sample. Repeated acquisition of sparse subsets enabled superresolution imaging in all three dimensions. The technique holds promise for improving 3D imaging even further, and is applicable to materials science as well as biological and biomedical studies