Impact of SYTO
9 and propidium iodide (PI) concentration on
growth rate and the detection efficiency of dead bacteria in M9 bacterial medium. (a) Inhibition of bacterial growth in the presence of 6 μM SYTO® 9 and 30 μM PI. PI alone showed a dose-dependent growth inhibition. At a concentration of 30 μM E. coli growth was inhibited, which was not detected when the PI concentration was reduced tenfold from 30 to 3 μM. Replication of E. coli in 3 μM PI containing M9 medium was confirmed by growth rate measurements from a starting culture of 50% live / 50% dead E. coli (inset) (b) PI fluorescence of E. coli cultivated in M9 medium containing different PI concentrations. 3 μM PI sufficiently stained dead bacteria in a 50% live / 50% dead E. coli mixture, while no significant background signal increase was detected for 3 μM PI compared to the background for a 100% live bacterial solution. In contrast, supplementing the medium with 30 μM PI resulted in a significant increase of PI stained bacteria from a 100% live starting culture indicating that this high concentration of the DNA stain interferes with bacterial viability. Background fluorescence of PI supplemented M9 medium was subtracted for each of the three PI concentrations, respectively. Error bars represent the standard error of the mean.