Supplementary Figure S1. Inhibition of fH binding to intact meningococci from all 3 variant fHbp families by polyanions. Purified factor H (1 µg) was incubated with increasing doses of either sucrose octasulfate (SOS) (left panel) or heparin (right panel), followed by addition to intact bacteria (H44/76 (variant 1 fHbp; upper panel), RM1090 (variant 2 fHbp; middle panel) and M1239 (variant 3 fHbp; lower panel). Bound fH was detected using mAb 90X followed by anti-mouse IgG-FITC. The positive control was bacteria incubated with fH alone (solid line) and the negative control (background binding) was bacteria incubated with the primary and secondary Abs (no fH added; light dotted line).
Supplementary Fig. S1 SOS Heparin H44/76 (variant 1) RM1090 (variant 2) M1239 (variant 3) control control fH fH fH+SOS 10mM fH+heparin 100 u/ml fH+SOS 40mM fH+heparin 400 u/ml
Supplementary Figure 2. Differential amounts of fH and FHL-1 binding to N. meningitidis and N. gonorrhoeae PorB.1A. N. meningitidis strain H44/76 (fHbp variant 1; left graph) and N. gonorrhoeae PorB.1A-expressing strain 15253 (right graph) were incubated with equimolar amounts (120 nM) of the full length purified fH and FHL-1. fH and FHL-1 bound to bacteria was detected using mAb 90X that is specific for fH SCR1 (and therefore also recognizes FHL-1), followed by anti-mouse IgG-FITC. Binding of the full length fH is shown by the shaded histograms, binding of FHL-1 by the solid lines and controls (no fH or FHL-1 added) by the broken lines. The x-axis represents fluorescence on a log10 scale and the y-axis the number of events on a linear scale.
Supplementary Fig. S2 N. meningitidis strain H4476 Variant 1 fHbp N. gonorrhoeae strain 15253 PorB.1A Isotype control Purified human factor H 18 µg/ml (~120 nM) Purified human FHL-1 5 mg/ml (~120 nM)