A simple method to prepare microorganisms for AFM analysis

Nowadays AFM becomes a more and more attractive method for microbiologists to investigate Microorganisms. The technique allows imaging over a broad magnitude scale and is not confined by the diffraction limit. Sometimes it is interesting to measure the dimensions of an organism. The other time the question is about surface properties of a cell. The scanning principle makes the AFM technique comparatively slow and the specimen has to be fixed on a flat surface during the scans. It is quite simple to dry the samples on a surface. Drying leads to a good immobilization but also to drying artifacts like denaturation of Proteins and shrinkage of the whole cell due to the loss of water. Therefore it is advantageous for most biological questions to do the imaging in liquids. Immobilization is not trivial as result of the heterogeneous surface properties of different micro organisms. Existing preparation methods are mostly utilizing coated surfaces or lithographicaly prepared surfaces. While lithography is not an option for everyone there is a variety of coatings available for instance poly-L-lysine or gelatine which work quite good with some microorganisms. A new method based on polyelectrolyte coated surfaces combined with centrifugal sedimentation shows promising results regarding the efficiency of immobilization. A variety of micro organisms were tested with the new method showing universality for many organisms. The samples were prepared with and without fixation. Of course fixation simplifies the imaging by enhancing the stability of the samples. But even unfixed Microorganisms can be imaged which opens the field for investigations in respect to cell division or other dynamic processes of living cells.