Principles Of Light Microscopy

For light microscopy, visible light is passed through the specimen and then through a series of lenses that reflect the light in a manner that results in magnification of the organisms present in the specimen (Figure 6-1). The total magnification achieved is the product of the lenses used.


In most light microscopes, the objective lens, which is closest to the specimen, magnifies objects lOOx (times) and the ocular lens, which is nearest the eye, magnifies 10x. Using these two lenses in combination, organisms in the specimen are magnified lOOOx their actual size when viewed through the ocular lens. Objective lenses of lower magnification are available so that those of lOx, 20x, and 40x magnification power can provide total magnifications of lOOx, 200x, and 400x, respectively. Magnification of lOOOx allows for the visualization of fungi, most parasites, and most bacteria but is not sufficient for observing viruses, which require magnification of 100,000x or more (see Electron Microscopy in this chapter).


To optimize visualization, other factors besides magnification must be considered. Resolution, defined as the extent to which detail in the magnified object is maintained, is also essential. Without it everything would be magnified as an indistinguishable blur.

BOX 6-1 Applications of Microscopy in Diagnostic _Microbiology

> Rapid preliminary organism identification by direct visualization in patient specimens » Rapid final identification of certain organisms by direct visualization in patient specimens 1 Detection of different organisms present in the same specimen

  • Detection of organisms not easily cultivated in the laboratory
  • Evaluation of patient specimens for the presence of cells Indicative of inflammation (i.e., phagocytes) or contamination (i.e., squamous epithelial cells)
  • Determination of an organism's clinical significance. Bacterial contaminants usually are not present in patient specimens at sufficiently high numbers (xlO5 cells/mL) to be seen by light microscopy

Provide preculture information about which organisms might be expected to grow so that appropriate cultivation techniques are used

• Determine which tests and methods should be used for identification and characterization of cultivated organisms Provide a method for investigating unusual or unexpected laboratory test results

Table 6-1 Microscopy for Diagnostic Microbiology

Organism Group

Bright-Field Microscopy

Fluorescence Microscopy

Bark-Field Microscopy

Electron Microscopy





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Bacterial Vaginosis Facts

Bacterial Vaginosis Facts

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