Comparison ofEscherichia colisurface attachment methods for single-cell,in vivomicroscopy

Abstract

For in vivo, single-cell imaging bacterial cells are commonly immobilised via physical confinement or surface attachment. Different surface attachment methods have been used both for atomic force and optical microscopy (including super resolution), and some have been reported to affect bacterial physiology. However, a systematic comparison of the effects these attachment methods have on the bacterial physiology is lacking. Here we present such a comparison for bacterium Escherichia coli, and assess the growth rate, size and intracellular pH of cells growing attached to different, commonly used, surfaces. We demonstrate that E. coli grow at the same rate, length and internal pH on all the tested surfaces when in the same growth medium. The result suggests that tested attachment methods can be used interchangeably when studying E. coli physiology.

A General Workflow for Characterization of Nernstian Dyes and Their Effects on Bacterial Physiology

“…Images were stabilized in x, y, and z positions using a bead attached to the coverslip and back-focal-plane interferometry (29,37). Cells grow attached to the poly-L-lysine surface with expected growth rates (given the medium), as previously reported (38) and seen in Videos S1 and S2.…”