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Fluorescence microscopy of fungi in native soil - Improvement by additional substances.
Christian Jensen and G. Lysek
Institute
of Systematic Botany and Plant Geography, Free University Berlin; Altensteinstr. 6, D-14195 Berlin.
First published by Microscopy and Anylysis in
September 1995
Issue No: 49 Pages: 7 - 9
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Introduction
The staining by fluorescein, applied in the form of fluorescein di-acetate, FDA, (SÖDERSTRÖM 1977; BAATH & SÖDERSTRÖM 1988), has allowed the
microscopic observation of predacious fungi in their native soil. This was possible by using the "hanging drop,' technique and after developing a procedure for cutting
the soil (JENSEN & LYSEK 1991; JENSEN 1994). The application of these methods allowed also the routine observation of these fungi during longer and
representative periods (SAXENA & LYSEK 1993); it also provided proofs that growth, development and predacious activities virtually resembled those known
from agar-cultures. In spite of this success, the method was not completely satisfying. The main disadvantage resulted from the FDA, which is not fluorescing
by itself, but only after uptake into living organisms and subsequent liberation of the fluorescein by hydrolytic cleavage (SÖDERSTRÖM 1977). As a consequence,
only those (parts of) living cells or hyphae show the fluorescence, which are actively metabolizing and thus have the hydrolytic capacity to transform the FDA
(CORREA et al. 1986). Other parts, especially older but still vital or dormant organs or structures, are not stained and hence not visualized by this otherwise
convenient method. Another problem hampering the broad usage of this method is the fast photo-fading of the fluorescein, especially at higher magnifications. It
limited the time available for observations and interfered heavily with the photographic documentation. Experiments thus were started, to use additional
staining and reacting substances. Among a number of chemicals few turned out to be valuable, but with their help the method could be improved considerably.
Materials and methods
Results (with figures)
Discussion
References
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