Abstract
•PES membrane used to immobilize of live B. laterosporus bacteria for AFM imaging.•At high resolution, the live bacteria displayed a certain pattern (“brain-like structure”).•Bacterial viability has no affect by the immobilization methods, as judged by the results of the growth-based enumeration of colonies.
One of the main obstacles to studying the surface ultrastructure of microbial cells by atomic force microscopy (AFM) is determining how to immobilize live cells on the AFM substrates. Each method has its own advantages and disadvantages. The aim of this study was to characterize a new simple and inexpensive method using two types of polyethersulfone (PES) membrane filters that differ in pore size (micropore and nanopore) to immobilize live and dead Brevibacillus laterosporus for AFM imaging. B. laterosporus was easily trapped by the microporous PES membrane, facilitating the successful AFM scanning of the bacterial surface ultrastructure. In addition, B. laterosporus strongly attached to the nanoporous membranes and withstood the pulling forces exerted by the AFM tip during scanning. These methods of immobilization did not affect the cell viability. The nanostructure and roughness of the bacterial surface were also observed for live, fixed, and air-dried cells. Live and dead bacteria displayed similar morphologies at low resolution, while at high resolution, live bacteria displayed a more convoluted surface ("brain-like structure").