Journal: Infection and Immunity

Article Title: Mechanisms of Decreased Susceptibility to β-Defensins by Treponema denticola

doi: 10.1128/iai.01718-06

Figure Lengend Snippet: FIG. 4. Quenching of a fluorescent dye in the presence of efflux pump inhibitors. A total of 1 108 cells/ml of T. denticola 35404 were incubated in 12 replicates at 37°C with 10 g/ml reserpine, 20 g/ml verapamil, 50 M sodium orthovanadate (NaoV), or the appropriate solvents. Then 10 l/well of a 10.25-g/ml acriflavine solution was added, and the fluorescence was determined immediately and at 2- to 3-min intervals. The percent quenching of acriflavine fluorescence compared to the fluorescence of the controls (solvent only) was cal- culated as follows: 100 ([average relative fluorescence units of wells with efflux inhibitor/average relative fluorescence units of wells with solvent only] 100). This experiment was conducted four times (12 replicates per condition), and the means and standard errors are shown.

Article Snippet: A total of 1 105 motile treponemes in 1 ml were added to triplicate tubes and incubated with 10 g/ml of human -defensin 2 (h D-2) or h D-3 (Peptides International, Lexington, KY) or 80 g/ml erythromycin (positive control for killing) at 37°C anaerobically for 0.5 to 4 h. In some experiments, efflux pump inhibitors, such as CCCP (final concentration, 35 M), reserpine (10 g/ml), verapamil (20 g/ml), or sodium orthovanadate (50 M), or equivalent amounts of their solvents were included in the killing assay mixture 10 min before addition of the defensin peptide.

Techniques: Incubation, Solvent