Apertureless near-field optical microscopy for fluorescence imaging

摘要

This thesis describes the development of a new type of optical microscope, an apertureless near-field microscope for fluorescence imaging (FANSOM). We have demonstrated that it is possible to use the probe of an atomic force microscope (AFM) to create a detectable modulation of the fluorescence of a nanometer-scale fluorescent object, and that it is possible to use that modulation to produce images by raster-scanning. Our results show that such a microscope is capable of a lateral resolution better than 20 nm.The processes that cause this modulation act for very small probe-sample separations, smaller than 20 nm. We have demonstrated FANSOM contrast using two completely different effects, one being a fluorescence inhibition, the other being a fluorescence enhancement generated by electric field enhancement. We have obtained contrast ratios of 1.90:1 using the fluorescence inhibition effect, and 5:1 using the fluorescence enhancement effect.Fluorescence can be inhibited by the proximity of a metallic probe. We have mapped the vertical profile of this phenomenon, and have shown that its shape is dependent upon the method of illumination. With evanescent illumination, the closest range of the interaction can exhibit some fluorescence enhancement that partially cancels the fluorescence inhibition effect.When a metallic or dielectric probe approaches a sample illuminated by an evanescent illumination field polarized parallel to the probe (i.e., vertically with respect to the surface), field enhancement occurs. A local fluorescence enhancement is detected as a result of the field enhancement. The range of this effect is less than 10 nm, located mostly under the AFM probe; the optical images acquired using this contrast mechanism exhibit a lateral resolution equal to or higher than the topographic resolution measured by AFM.The design of the data acquisition system allows us to obtain precise mappings of signal intensity to probe--sample separation, more precise than have previously been achieved in an apertureless near-field microscopy system. It has allowed us to obtain repeatable approach curves that differ clearly from one another when the type of illumination or probe is changed.