Figure 4
From: Antibacterial activity of ceramide and ceramide analogs against pathogenic Neisseria
(A) Schematic of bacteria treated with ceramide analogs, followed by click reaction with DCBO-Sulfo-Cy5. (B) Azido-modified ceramides are rapidly incorporated into N. meningitidis. Bacteria were treated with 5 µM of α-azido-C6-ceramide (α-C6-cer), ω-azido-C6-ceramide (ω-C6-cer), α-azido-C16-ceramide (α-C16-cer) and ω-azido-C16-ceramide (ω-C16-cer), or DBCO-sulfo-Cy5 (DBCO Cy5) for indicated time points, washed, and ceramide incorporation levels were detected after 15 min DBCO-sulfo-Cy5-clicking and flow cytometry analysis. Three independent experiments with SD are shown. *, **, ***, ****P < 0.05, 0.01, 0.001, 0.0001 in one-way ANOVA and Tukey post hoc test relative to dye control (DCBO-sulfo-Cy5). (C) LSM images of N. meningitidis after incorporation of (A) α–azido-C6-ceramide, (B) ω–azido-C6-ceramide, (C) α–azido-C16-ceramide, and (D) ω–azido-C16-ceramide and clicking with DBCO-Sulfo-Cy5 by copper-free biorthogonal click chemistry. For visualization the contrast was adjusted for each image individually. Scale bar, 10 µm.
