Scanning Microscopy


A common problem with electron micrographs of biological objects is that fine details are usually faint and, moreover, tend to be obscured by background noise from stain and support film. Filtering is a useful way to improve the signal-to-noise ratio and is particularly important when trying to detect small changes (conformational or due to labelling) or when examining frozen hydrated objects, where the statistical definition of the image has been reduced because of the low-dose conditions needed to prevent radiation damage. Filtering can also be used to address the superposition effects that result from the large depth of focus of electron microscopes and is particularly effective when dealing with the Moiré patterns produced by overlapping regular layers as found, for example, in helices or tubes. The alteration of the image by the non-uniform phase contrast transfer function of the microscope can also be compensated for by using image processing in conjunction with electron diffraction. Finally, the essentially two-dimensional nature of the information can be extended to three-dimensions by combining views from different orientations. Fourier-based methods are particularly effective when dealing with regular objects, such as crystals, helices and shell structures such as icosahedral viruses.

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