Optical microscopy, resolution limitation

In microscopy, spatial resolution is the ability to view two closely spaced objects as distinct particles. The maximum spatial resolution in a conventionally designed far-field microscope is wavelength limited. The major drawbacks of optical microscopy, namely limited resolution, poor contrast and restricted depth... 

The spatial resolution of conventional optical microscopy is limited by the diffraction limit of light consequently, two objects that are close to each other cannot be resolved in the fluorescence image. The minimum resolvable distance. Ax, is given as follows ... 

While the spatial resolution in classical microscopy is limited to approximately X/2, where X is the optical wavelength (tlie so-called Abbe Limit [194], -0.2 pm with visible light), SNOM breaks through this barrier by monitoring the evanescent waves (of high spatial frequency) which arise following interaction with an... 

The resolution or "resolving power" of a light microscope is usually specified as the minimum distance between two lines or points in the imaged object, at which they will be perceived as separated by the observer. The Rayleigh criterion [42] is extensively used in optical microscopy for determining the resolution of light microscopes. It imposes a resolution limit. The criterion is satisfied, when the centre of the Airy disc for the first object occurs at the first minimum of the Airy disc of the second. This minimum distance r can then be calculated by Equation (3). 

The resolution of a conventional microcope is limited by the classical phenomena of interference and diffraction. The limit is approximately X/2, X being the wavelength. This limit can be overcome by using a sub-wavelength light source and by placing the sample very close to this source (i.e. in the near field). The relevant domain is near-field optics (as opposed to far-field conventional optics), which has been applied to microscopy, spectroscopy and optical sensors. In particular, nearfield scanning optical microscopy (N SOM) has proved to be a powerful tool in physical, chemical and life sciences (Dunn, 1999). 

Optical microscopy is often the first step in surface analysis, since it is fast and easy to perform. It can be an aid in selecting the area of interest on a sample for further analysis with more complex methods. The application of classical optical microscopy to surface science is, however, limited because the maximum lateral resolution is in the order of the optical wavelength ( 500 nm). For opaque solids, the light penetrates into the material, giving optical microscopy a poor surface sensitivity. In addition, the depth of field is limited which calls for flat, polished surfaces or allows only plane sections of the sample to be viewed. 

Three major advancements in resolution have occurred since Hookes s discovery of the optical microscope in 1665 [46]. In 1873, Ernst Abbe established fundamental criteria for the resolution limit in optical microscopy [47], which did not exceed the range of a couple of 100 nanometers even after the introduction of the confocal optical microscope [43,48]. The invention of the transmission electron microscope by Ernst Ruska in 1933 extended the resolution of microscopes to the nanometer scale [49]. Finally, scanning tunnelling microscopy introduced, by Binnig and Rohrer in 1981, made a breakthrough when atomic... 

SNOM combines the optical contrast with a high lateral resolution of SPMs [55,56]. Scanning a surface with a sharp optical fibre tip within the range of the optical near field makes it possible to overcome the optical diffraction limit that restricts the resolution of conventional optical microscopy. Moreover, the SNOM probe operates at a finite distance from the surface, so that damage and distortion of delicate samples can be eliminated. The drawback of SNOM compared to other SPM methods is its relatively low resolution - around tens of nanometers [62,63]. 

Particle sizes as measured by optical microscopy are likely to be in serious error for diameters less than c. 2 pm, although the limit of resolution is some ten times better than this (see Table 3.1). 

Two techniques for overcoming the limitations of optical microscopy are of particular value in the study of colloidal systems. They are electron microscopy36-37, in which the limit of resolution is greatly extended, and dark-field microscopy, in which the minimum observable contrast is greatly reduced. 

Decreasing the nanochannel width thus leads to qualitatively new and counterintuitive behavior that can be exploited for molecular separations. Because details of the flow profiles in individual nanochannels are below the resolution limit of optical microscopy, only the average velocities of dye fronts can be monitored. Significant improvements in the lateral resolution of analytical imaging methods are required to study the transport of molecules in an individual channel. 

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