The attractive feature of optical microscopy is that it is so easy: samples can be analyzed in air or water, the images are in natural color with magnifications of up to one hundred to one thousand times, and modern semiconductor electronics with charge-coupled devices (CCD) allow image processing.

The optical microscope should dominate the field. It doensn't. The scanning electron microscope (SEM) is microscope of choice because of its depth of focus and resolving capability. Examination of Fig. 2 shows a striking contrast between an a) optical and b) SEM viewgraph of a radiolarian at the same magnification.

Figure 2. (Taken from J.I. Goldstein et al., eds., Scanning Electron Microscopy and X-Ray Microanalysis, (Plenum Press,NY,1980).)

In the optical micrograph taken at high resolution only a section of the radiolarian is in sharp focus. In the lower SEM image the whole specimen is in focus.

For the optical microscope, the depth of focus is the distance above and below the image plane over which the image appears in focus. As the magnification increases in the optical microscope the depth of focus decreases.

The three-dimensional appearance of the specimen image is a direct result of the large depth of field of the SEM. It is these large depth of fields in the SEM that is the most attractive feature of the scanning electron microscope. This field arises because of the method in which the data is obtained with a fine electron beam scanned over the surface and with the detected secondary electrons forming an image on the "TV"-like monitor.

The other feature of scanning electron microscopy is resolving power which is the smallest detail that a microscope can resolve, or "see". The resolving power of electron microscopes is orders of magnitude better than that of an optical microscope because the wavelength of the probing beam is orders of magnitude smaller.

The limit on what size can be resolved -- irrespective of instrument type -- is set by the wavelength. The wavelength of the visible light used in optical microscopes is between 400 and 700 nanometers (nm). The resolving powers of high-quality light microscopes are limited by the wavelength of imaging light to about 200 nanometers (0.2 microns, 0.2meter). Scanning electron microscopy uses electrons with energies of a few thousand electron volts (eV), energies a thousand times greater than that of visible light (2 to 3 eV). The wavelength is given by h / momentum where h is Planck's constant. For 3600 electron-volt electrons, the wavelength is 0.02 nanometers. The resolving powers of ordinary electron microscopes is 1 nanometer (above the limit because construction details determine resolving power), a value which can be pushed to 0.1 nanometer.

Depth of focus and resolving power draws one to the SEM and operating maintenance as well as vacuum requirements drives one away. Electrons are light-weight (1/1836 the mass of the proton) and are scattered or absorbed in air. The sample chamber must be in vacuum which limits the sample size to a few centimeters on edge. Electrons carry charge (e = 1.6 x 10^-19 Coulomb). The samples must be covered with a conducting coating. Finally, the SEM is expensive and requires maintenance.

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