Figure 1
MAC formation and NHS-mediated reduction in the growth of S. aureus is dependent on a complete terminal complement pathway. (a) S. aureus was incubated with 50% NHS (top) or media (bottom) for 8 h and then visualized by scanning electron microscopy (SEM). (b) S. aureus was incubated with 50% NHS (top), 50% C5-depleted serum (middle) or media (bottom) for 1 h. Complement proteins indicated at the top of each column were detected using antibodies specific for C1q (far left), C3 (center) or C5b9 (far right) followed by an Alexa Fluor 488 conjugated secondary (green). Bacteria were stained with DAPI (blue) and then imaged using a Zeiss LSM780 confocal microscope. (c) The effect of human serum on growth of S. aureus was measured using serum killing assays. S. aureus was incubated with 50% of the indicated sera or medium for 24 h. After incubation, bacteria were enumerated by serial dilution and plating. Results are plotted as mean with standard deviation. *P < 0.05, ***P < 0.001, ****P < 0.0001, one-way ANOVA with Dunnett’s test showing significance compared to the NHS sample. (d) S. aureus was incubated with 50% NHS, 50% C1q-depleted serum or 50% C5-depleted serum for 0, 10, and 24 h at 37 °C. Uptake of the viability dye propidium iodide (PI) was assessed by measuring fluorescence (excitation 533 nm, emission 617 nm). Results are plotted as mean with standard deviation. *P < 0.05, ****P < 0.001, two-way ANOVA showing values significantly different than NHS.
