Bacterial traps and advances in nanobiotechnology for atomic force microscopy

The atomic force microscope (AFM) allows the analysis of living microorganisms in physiological conditions on the nanometer scale. The observation of bacteria in physiological aqueous medium necessitates a robust immobilization of the bacterium to the surface, in order to withstand the lateral forces exerted by the AFM cantilever tip during scanning. Different immobilization techniques for AFM analysis of bacteria in aqueous media have been developed hitherto, however the immobilization techniques were dependent on the bacterial species and/or the aqueous imaging medium.

We propose a robust bacterial immobilization method allowing bacterial species and medium independent analysis. We demonstrate the immobilization and AFM analysis of different bacterial species such as gram-positive and -negative, motile and non-motile, and rod-shaped, ovococcal, and crescent bacteria. The developed bacterial traps were used together with Escherichia coli, Bacillus subtilis, Caulobacter crescentus, Streptococcus pneumoniae, and Acidiphilium cryptum bacteria in their corresponding physiological aqueous medium.

The developed microfluidic device allows simultaneous fluorescence and atomic force microscopy of bacteria. Moreover, we developed two different cleanroom microfabrication techniques for the bacterial traps. We thus fabricated nanotailored bacterial traps, allowing the immobilization of rod-shaped bacteria along their longitudinal axis as well as by the bacterial poles.

Furthermore, we discuss the nanomechanical analysis of suspended silicon nanowires and hydrogels using the AFM. In the final part of the thesis, we explain the microfabrication method for AFM cantilevers with a low quality factor and elucidate hard tip integration into the developed multilayer AFM cantilevers.