Graf B, Göbel U B, Adam T
Institut für Mikrobiologie und Hygiene, Medizinische Fakultät der Humboldt-Universität, Universitätsklinikum Charité, Berlin, Deutschland.
Mycoses. 1998;41 Suppl 1:39-46. doi: 10.1111/j.1439-0507.1998.tb00582.x.
Fluorescence microscopy is an important method in mycology. It is a common procedure used in immunology or histology and more recently in modern techniques of molecular biology like in-situ hybridization. Since several molds and yeasts show autofluorescence, an interference of this phenomenon with the detection method cannot be excluded. Therefore, we studied autofluorescence in fungi in more detail, in particular with respect to the dependence of this phenomenon from growth conditions, fixing method or mounting medium used. Here we show that moulds cultivated in a liquid medium are strongly autofluorescent which could be considerably reduced by repetitive washing. In moulds, we did not find important differences in autofluorescence levels with the three fixing methods under study. However, this finding cannot be generalized. Thus, in the yeast Candida albicans we found the autofluorescence pattern being largely dependent from the fixing method and the excitation wave length, respectively. In particular, with green excitation we could show that aceton fixation resulted in strong fluorescence of individual cells within a vast population of cells showing little or no autofluorescence. In addition, we could demonstrate that mounting media are able to strongly modify autofluorescence in fungi. Using digital image acquisition with a cooled CCD camera we were able to quantify the influence of different mounting media on fluorescence intensities of Aspergillus fumigatus.
