5 - Recent Advances in Optical Microscopy and its Application to the Characterization of Polymers

Abstract

An optical microscope is perhaps one of the most used imaging instruments in a variety of disciplines to ‘see’ things at high magnification. One of the limitations of traditional optical microscopy techniques (far-field techniques) is its diffraction-limited resolution.1 Despite the limited resolution, its widespread use as an imaging instrument is due to its simplicity and a unique variety of available contrast mechanisms. Examples of such contrast mechanisms are absorption, polarization, phase contrast, dark-field, and fluorescence, to name just a few. Fluorescence microscopy in particular has revolutionized cell biology, allowing one to obtain images of cells and organelles in exquisite detail. Such advances have been made by combining lasers, electronic cameras, and digital image analysis. In view of the advantages provided by various contrast mechanisms and the ease of use, efforts to push the resolution (both spatial and axial) of the optical microscope well beyond that of the diffraction limit have produced a number of very useful techniques, such as confocal microscopy,2 standing-wave fluorescence microscopy (SWFM)3 (where one is able to visualize structure in 3D by optical sectioning of the object being viewed), solid immersion microscopy (SIM),4 photon tunneling microscopy (PTM),5, 6 and, at a higher spatial resolution, near-field optical microscopy (NSOM).7–10 All of these techniques, by virtue of using light at visible wavelengths, provide information that is not accessible by other high-resolution techniques such as electron microscopy and the recent family of probe microscopes (scanning tunneling microscopy and force microscopy).11–14 We will not dwell on such forms of microscopy and will refer the reader to excellent reviews and books that have been written.14