Effect of glutaraldehyde fixation on bacterial cells observed by atomic force microscopy

Abstract

Atomic force microscopy (AFM) is a promising microscopy technique that can provide high-resolution images of bacterial cells without fixation. Three species of bacteria, Xanthomonas campestris, Pseudomonas syringae, and Bacillus subtilis, were used in this study. AFM images were obtained from unfixed and glutaraldehyde-fixed cells, and cell height was measured. The mean height of bacterial cells prepared by fixation was higher than that of those prepared by nonfixation. However, the height changes were different between Gram-negative and Gram-positive bacteria: the mean height of two fixed Gram-negative bacteria, X. campestris and P. syringae, increased by 112.31 and 84.08%, respectively, whereas Gram-positive bacterium, B. subtilis, increased only by 38.79%. The results above indicated that glutaraldehyde fixation could affect the measured height of cells imaged by AFM; further more, the effect of glutaraldehyde fixation on the measured height of Gram-negative bacterial cells imaged by AFM seemed much more than on that of Gram-positive bacterial cells.

Cell envelope disruption of E. coli Exposed to ϵ‐polylysine by FESEM and TEM technology

“…), which seriously hinders the observation of flexible and fine ultrastructure and even leads to artifacts (Liu et al . ).…”

“…FESEM is very well suited to provide a quick overview of sample and can capture the surface details of specimens, but the information on the interior of cell can be visualized only by TEM after sectioning cell and staining (Milillo et al 2011). The key factor of the desired and lossless image is specimen processing for electron microscopy, conventional specimen preparation methods involve more steps for example, chemical fixation, staining and dehydration by replacement (Diestra et al 2005;Azad et al 2011), and undergo more centrifugations (Anderson et al 2004;Bouhdid et al 2009), which seriously hinders the observation of flexible and fine ultrastructure and even leads to artifacts (Liu et al 2012).…”

Glutaraldehyde fixation preserves the trend of elasticity alterations for endothelial cells exposed to TNF‐α

“…The fixation of cells for AFM applications has been thoroughly studied [Braet et al, ; Weyn et al, ; Moloney et al, ; Mc Namee et al, ; Méndez‐Vilas et al, ; Codan et al, ], however it mainly reports on the improvement of surface details imaging and the cell stiffening (over an order of magnitude). The post‐fixation cell stiffening is a well‐known and widely described effect [Weyn et al, ; Braet et al, ; Hutter et al, ; Liu et al, ]. Moreover, there has been an indication that after fixation the elastic properties of cells are more unified over the whole cell surface [Yamane et al, ].…”

Microfluidic bacterial traps for simultaneous fluorescence and atomic force microscopy

“…The optimal coating depends strongly on the bacterial species and strain, as well as the surrounding liquid imaging medium. Nevertheless, the bacterial envelope properties are altered when the bacterium is attached to a chemically modified surface, which can additionally trigger bacterial responses and compromise the analysis of the bacterial viability [16,17,44,45].…”