Microfluidic bacterial traps for simultaneous fluorescence and atomic force microscopy

The atomic force microscope has become an established research tool for imaging microorganisms with unprecedented resolution. However, its use in microbiology has been limited by the difficulty of proper bacterial immobilization. Here, we have developed a microfluidic device that solves the issue of bacterial immobilization for atomic force microscopy under physiological conditions. Our device is able to rapidly immobilize bacteria in well-defined positions and subsequently release the cells for quick sample exchange. The developed device also allows simultaneous fluorescence analysis to assess the bacterial viability during atomic force microscope imaging. We demonstrated the potential of our approach for the immobilization of rod-shaped Escherichia coli and Bacillus subtilis. Using our device, we observed buffer-dependent morphological changes of the bacterial envelope mediated by the antimicrobial peptide CM15. Our approach to bacterial immobilization makes sample preparation much simpler and more reliable, thereby accelerating atomic force microscopy studies at the single-cell level.