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High-resolution Visualization of the Microbial Glycocalyx with Low-voltage Scanning Electron Microscopy: Dependence on Cationic Dyes

2004; SAGE Publishing; Volume: 52; Issue: 11 Linguagem: Inglês

10.1369/jhc.4a6428.2004

1551-5044

Stanley L. Erlandsen, Christopher J. Kristich, Gary M. Dunny, Carol L. Wells,

Minerals Flotation and Separation Techniques

The microbial glycocalyx is composed of a variety of polyanionic exopolysaccharides and plays important roles in microbial attachment to different substrata and to other cells. Here we report the successful use of low-voltage scanning electron microscopy (LVSEM) to visualize the glycocalyx in two microbial models (Klebsiella pneumoniae and Enterococcus faecalis biofilms) at high resolution, and also the dependence on fixation containing polycationic dyes for its visualization. Fixation in a paraformaldehyde-glutaraldehyde cocktail without cationic dyes was inadequate for visualizing the glycocalyx, whereas addition of various dyes (alcian blue, safranin, and ruthenium red) to the aldehyde cocktail appeared necessary for stabilization. The cationic dyes varied in size, shape, and charge density, and these factors appeared responsible for different phenotypic appearances of the glycocalyx with each dye. These results suggest that aldehyde fixation with cationic dyes for high-resolution LVSEM will be a useful tool for investigation of microbial biofilms as well as investigation of the extent and role of the glycocalyx in microbial attachment to surfaces.