Staining of Heterotrophic Protists for Visualization via Epifluorescence Microscopy

Studies of the in situ abundance and activity of heterotrophic protists, especially cells <20 µm in size, were hampered due to inadequate methods of enumeration of this group of microbes until epifluorescence microscopical techniques began to be applied to the problem in the early 1980s. Since photosynthetic pigments fluoresce, epifluorescence microscopy may also be used to distinguish photo trop hie protists that can be abundant in the nanoplankton size class. The autofluorescence of photosynthetic pigments varies from yellow to red and is dependent upon the type of pigment and the filter set employed. A brief description of fluorochrome staining methods currently used to visualize heterotrophic protists is given. The great advantage of epifluorescence microscopy for counting protistan (and prokaryotic) populations relative to most other techniques is the relative ease of the method. Critical ultrastructural features cannot be discerned with this method, and electron microscopy and the observation of live specimens remain important methods for identifying small protists.