<i>N. meningitidis</i> and <i>N. gonorrhoeae</i> isolates.

<p><i>N. meningitidis</i> and <i>N. gonorrhoeae</i> isolates.</p

Immunogenicity of Ghfp.

<p>(<b>A</b>) Detection of fHbp variants in whole cell lysates of <i>N. meningitidis</i> by Western blot analysis using anti-Ghfp serum. Recognition of recombinant V1 (<b>B</b>), and V2and V3 (<b>C</b>) fHbps by anti-Ghfp serum by ELISA. (<b>D</b>) SBA responses for anti-Ghfp serum.</p

Immune responses against isogenic <i>N. meningitidis</i> strains.

<p>(<b>A</b>) Western blot analysis of whole cell lysates of fHbp expressed by isogenic MC58Δ<i>fhbp</i> strains detected by anti-Ghfp serum. (<b>B</b>) Surface expression of different fHbps in the isogenic MC58Δ<i>fhbp</i> strains detected by flow cytometry. Graph shows the mean ± SEM of three separate experiments. (<b>C</b>) Representative corresponding flow cytometry overlay of MC58 (grey hatched area), MC58Δ<i>fhbp</i> and MC58Δ<i>fhbp</i>+<i>fhbp</i> V1.1 detected by anti-Ghfp by flow cytometry. (<b>D</b>) SBA responses against isogenic MC58 strains for anti-Ghfp, fHbp V1.1, and fHbp V3.45 serum using rabbit complement; NK, no killing. (<b>E</b>) SBA responses against isogenic H44/76 strains with anti-Ghfp serum using rabbit complement.</p

fH binding to modified fHbp V3.45.

<p>(<b>A</b>) Analysis of fH binding to modified V3.45 fHbp by far Western using normal human serum as the source of fH. Molecular mass is shown in kDa. (B) SPR values of fH_6–7 binding to wild type and modified V3.45 fHbp; NBD, no binding detected. (<b>C</b>) Detection of full length fH (5 nM) binding to wild-type and modified V3.45 fHbp by ELISA. Data represents the mean ± SEM of three different experiments.</p

Ghfp is not expressed on the surface of <i>N. gonorrhoeae</i>.

<p>(<b>A</b>) Western blot analysis of Ghfp expression by <i>N. gonorrhoeae</i> strains F62, F62Δ<i>ghfp</i> and FA1090, and fHbp V3.28 expressed by <i>N. meningitidis</i> strains M1239 and M1239Δ<i>fhbp</i> using anti-Ghfp serum. (<b>B</b>) Western blot analysis of Ghfp expressed by a panel of clinical <i>N. gonorrhoeae</i> isolates (GC1-11, inclusive). Surface expression of Ghfp (<b>C</b>) and fHbp V3.28 (<b>D</b>) was assessed by flow cytometry analysis using anti-Ghfp serum. Error bars, SEM of three separate experiments; ** p<0.05 and NS (not significant, Student's <i>t</i>-test). Representative flow cytometry overlays are shown below the graphs. Bacteria incubated without anti-Ghfp serum are shown as the grey filled areas. (<b>E</b>) Bacteria were exposed to proteinase K (3 ng/ml and serial three-fold dilutions) and the effect on proteins (shown) determined by Western blot analysis using anti-Ghfp, anti-RecA and anti- α-Lst serum.</p

Oligonucleotides used in this study.

<p>Oligonucleotides used in this study.</p

fH binding capacity of Ghfp.

<p>(<b>A-C</b>) fH binding to wild type and modified Ghfp and fHbp V3.45 was assessed by far Western analysis using normal human serum as the source of fH. Western blots are representatives of three separate experiments. Molecular mass is shown in kDa. (<b>D</b>) Typical equilibrium fit for binding of fH_6–7 to Ghfp^M4–5. (<b>E</b>) SPR was performed with Ghfp, Ghfp^M4 (R288H), Ghfp^M5 (D318G) and Ghfp^M4–5 (R288H/D318G); NBD, no binding detected. (<b>F</b>) Detection of full length fH (5 nM) binding to wild-type and modified Ghfp by ELISA. Error bars, SEM of three separate experiments; *** p<0.01 and * p<0.1 (Student's <i>t</i>-test). (<b>G</b>) Cartoon presentation of the predicted structure of Ghfp (Yellow) and fH (Blue). Residues M1 (R176), M2 (D199) and M3 (D212) are shown in green while those amino acids that are important for fH binding, M4 (R288) and M5 (D318G), are shown in red.</p

Binding of C3 (A and C) and MAC (B and D) to bacteria detected by FACS analysis after incubation in immune human serum pooled from 10 donors

Results are shown as the MFI (calculated as geometric mean multiplied by the percentage of gated cells), and the error bars show the SEM. In B and D, strains were incubated with serum in the presence of magnesium and EGTA to block the CP and LP (-CP/LP). Error bars show the SEM of assays performed in triplicate. There was significantly less binding of complement factors to R3 and S3∷R compared with S3. P < 0.01 in all assays with the Student's test.<p><b>Copyright information:</b></p><p>Taken from "A generic mechanism in for enhanced resistance against bactericidal antibodies"</p><p></p><p>The Journal of Experimental Medicine 2008;205(6):1423-1434.</p><p>Published online 9 Jun 2008</p><p>PMCID:PMC2413038.</p><p></p

Capsular material was detected around the surface of the sensitive strain S3 (A and B, arrows) but not the corresponding mutant (C and D)

Strains R3 and S3∷R (E and F; and G and H, respectively) expressed larger amounts of capsule, which formed large extracellular aggregates in places. Bars, 400 nm.<p><b>Copyright information:</b></p><p>Taken from "A generic mechanism in for enhanced resistance against bactericidal antibodies"</p><p></p><p>The Journal of Experimental Medicine 2008;205(6):1423-1434.</p><p>Published online 9 Jun 2008</p><p>PMCID:PMC2413038.</p><p></p

(A) SBA titers using sera from eight vaccinees against the R strains and the control strain, C11

All sera had SBA titers of <p><b>Copyright information:</b></p><p>Taken from "A generic mechanism in for enhanced resistance against bactericidal antibodies"</p><p></p><p>The Journal of Experimental Medicine 2008;205(6):1423-1434.</p><p>Published online 9 Jun 2008</p><p>PMCID:PMC2413038.</p><p></p