Flow cytometry for bacteria: enabling metabolic engineering, synthetic biology and the elucidation of complex phenotypes

Role of Autofluorescence in Flow Cytometric Analysis of Escherichia coli Treated with Bactericidal Antibiotics

“…Early experiments by Steen et al showed how effects of antibiotics on bacterial cells can be assessed with flow cytometry (4-7). More recently, Tracy et al demonstrated the potential and the increased importance of flow cytometry in microbiology (1). In line with this, a recent proposal for a common mechanism of cell death induced by bactericidal antibiotics is based on data obtained with flow cytometry (8)(9)(10).…”

“…ith flow cytometry, it is possible to measure several parameters simultaneously and details about physical and biological properties of single particles in a population are obtained (1,2). It is an established technique for studies in cell and developmental biology.…”

“…Still, with bacteria the potential of this technology has not been fully exploited (1,3). The main problems are small cell size, the low per-cell content of macromolecules, and the selection of the most suitable fluorescent dyes that had been originally developed for mammalian cells (1)(2)(3).…”

“…The main problems are small cell size, the low per-cell content of macromolecules, and the selection of the most suitable fluorescent dyes that had been originally developed for mammalian cells (1)(2)(3). For example, prokaryotes do not share the permeability or the uptake kinetics of eukaryotic cells, and bacterial efflux mechanisms efficiently pump out many fluorochromes (1,2).…”

Plasmonic imaging of protein interactions with single bacterial cells

“…The data generated with these bulk assays are averaged over many bacteria, which wash out important variability or heterogeneity of different bacterial cells. Various imaging techniques, such as fluorescence (Cywes--Bentley et al, 2013;Sochacki et al, 2011), AFM (Fantner et al, 2010;Lower et al, 2011) and transmission electron microscopy (TEM) (Cywes--Bentley et al, 2013), and non--imaging microfluidics techniques, such as flow cytometry (Tracy et al, 2010) and micro electrophoresis (van der Mei and Busscher, 2012), have been used to study bacterial surfaces. These techniques have contributed to the understanding of bacteria, but each has disadvantages.…”