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000819825 1001_ $$0P:(DE-Juel1)156207$$aKrämer, Christina$$b0$$ufzj
000819825 245__ $$aTime-resolved, single-cell analysis of induced and programmed cell death via non-invasive propidium iodide and counterstain perfusion
000819825 260__ $$aLondon$$bNature Publishing Group$$c2016
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000819825 520__ $$aConventional propidium iodide (PI) staining requires the execution of multiple steps prior to analysis, potentially affecting assay results as well as cell vitality. In this study, this multistep analysis method has been transformed into a single-step, non-toxic, real-time method via live-cell imaging during perfusion with 0.1 μM PI inside a microfluidic cultivation device. Dynamic PI staining was an effective live/dead analytical tool and demonstrated consistent results for single-cell death initiated by direct or indirect triggers. Application of this method for the first time revealed the apparent antibiotic tolerance of wild-type Corynebacterium glutamicum cells, as indicated by the conversion of violet fluorogenic calcein acetoxymethyl ester (CvAM). Additional implementation of this method provided insight into the induced cell lysis of Escherichia coli cells expressing a lytic toxin-antitoxin module, providing evidence for non-lytic cell death and cell resistance to toxin production. Finally, our dynamic PI staining method distinguished necrotic-like and apoptotic-like cell death phenotypes in Saccharomyces cerevisiae among predisposed descendants of nutrient-deprived ancestor cells using PO-PRO-1 or green fluorogenic calcein acetoxymethyl ester (CgAM) as counterstains. The combination of single-cell cultivation, fluorescent time-lapse imaging, and PI perfusion facilitates spatiotemporally resolved observations that deliver new insights into the dynamics of cellular behaviour.
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000819825 7001_ $$0P:(DE-Juel1)129076$$aWiechert, Wolfgang$$b1
000819825 7001_ $$0P:(DE-Juel1)140195$$aKohlheyer, Dietrich$$b2$$eCorresponding author
000819825 773__ $$0PERI:(DE-600)2615211-3$$a10.1038/srep32104$$gVol. 6, p. 32104 -$$p32104$$tScientific reports$$v6$$x2045-2322$$y2016
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