Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse-2

<p><b>Copyright information:</b></p><p>Taken from "Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse"</p><p>http://www.virologyj.com/content/4/1/118</p><p>Virology Journal 2007;4():118-118.</p><p>Published online 31 Oct 2007</p><p>PMCID:PMC2186315.</p><p></p>e) by i.n. or s.c. route. Mice were bled 3 weeks after the third immunization. Pooled plasma samples (3–5 mice/pool) were tested by ELISA for M2e(pep)- (dots) and M2e(pep-nat)-specific (circles) Ab titers. Horizontal bars indicate GMTs within each set. Data from four independent vaccination experiments are shown. B, C, D. 7–10 days after the third vaccination, mice were challenged by i.n. inoculation of 5 μl X31 (1000 TCID/mouse). Five days later, nose, trachea and lung were tested for virus titer. Each symbol indicates the virus titer of an individual mouse. Horizontal bars indicate the GMT within each vaccination set. Dashed (top) and stipulated (bottom) horizontal lines indicate the mean virus titer of control mice and threshold of virus detection, respectively. Tissues with undetectable virus were assumed to be virus free. Data were analyzed by non-parametric ANOVA and Dunn's Multiple Comparison post test. M2e-MAP vaccination groups with statistically significant reduction in virus titer compared to the control group are indicated by asterisks right above the group and statistical differences between M2e-MAP vaccination groups by asterisks above two-sided arrows: p < 0.05 (*), p < 0.01 (**), p < 0.001 (***)

Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse-8

<p><b>Copyright information:</b></p><p>Taken from "Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse"</p><p>http://www.virologyj.com/content/4/1/118</p><p>Virology Journal 2007;4():118-118.</p><p>Published online 31 Oct 2007</p><p>PMCID:PMC2186315.</p><p></p>M2 (circles) as described in the method section, using the same reagents and incubation times for each assay. The mean OD (± SEM) above background of six replicates at each Ab dilution are shown. The three sigmoidal titration curves have similar EC50 values (-9.3 vs G40d, -9.5 vs Cys-M2e, -9.2 vs HeLa-M2). To further demonstrate the similarity between the three titration curves, OD values measured against HeLa-M2 were multiplied by 1.65 to generated the stipulated curve. A representative assay is shown. B. Pooled plasma samples (5 mice/group), obtained 3 wks after second (left column) and third (right column) immunization, were tested by ELISA for M2e-MAP- (squares), M2e(pep)- (triangles) and M2e(pep-nat)-specific (circles) Ab titers as described in the method section. The mice had been immunized with 3 μg M2e-MAP G40d and adjuvants by i.n. or s.c. routes as indicated below the x axis. Each symbol shows the mean serum Ab concentration determined in each sample by 2–3 independent assays. Data from a single vaccination experiment are shown. C. The fraction of M2e(pep-nat)-specific Abs is expressed as percent of the M2e(pep)-specific Ab concentration within each sample. Each dot indicates the % of anti-M2e(pep-nat) per group of 3–5 mice immunized by one of the protocols indicated below the x axis. In most groups, samples from secondary and tertiary responses were tested, and the mean % of these is shown. Horizontal bars indicate the geometric means within a vaccination protocol. Data from 12 independent vaccination experiments are shown. Groups immunized by different protocols did not differ significantly (ANOVA) with regards to percentage of anti-M2e(pep-nat)-specific Abs

Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse-1

<p><b>Copyright information:</b></p><p>Taken from "Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse"</p><p>http://www.virologyj.com/content/4/1/118</p><p>Virology Journal 2007;4():118-118.</p><p>Published online 31 Oct 2007</p><p>PMCID:PMC2186315.</p><p></p>M2 (circles) as described in the method section, using the same reagents and incubation times for each assay. The mean OD (± SEM) above background of six replicates at each Ab dilution are shown. The three sigmoidal titration curves have similar EC50 values (-9.3 vs G40d, -9.5 vs Cys-M2e, -9.2 vs HeLa-M2). To further demonstrate the similarity between the three titration curves, OD values measured against HeLa-M2 were multiplied by 1.65 to generated the stipulated curve. A representative assay is shown. B. Pooled plasma samples (5 mice/group), obtained 3 wks after second (left column) and third (right column) immunization, were tested by ELISA for M2e-MAP- (squares), M2e(pep)- (triangles) and M2e(pep-nat)-specific (circles) Ab titers as described in the method section. The mice had been immunized with 3 μg M2e-MAP G40d and adjuvants by i.n. or s.c. routes as indicated below the x axis. Each symbol shows the mean serum Ab concentration determined in each sample by 2–3 independent assays. Data from a single vaccination experiment are shown. C. The fraction of M2e(pep-nat)-specific Abs is expressed as percent of the M2e(pep)-specific Ab concentration within each sample. Each dot indicates the % of anti-M2e(pep-nat) per group of 3–5 mice immunized by one of the protocols indicated below the x axis. In most groups, samples from secondary and tertiary responses were tested, and the mean % of these is shown. Horizontal bars indicate the geometric means within a vaccination protocol. Data from 12 independent vaccination experiments are shown. Groups immunized by different protocols did not differ significantly (ANOVA) with regards to percentage of anti-M2e(pep-nat)-specific Abs

Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse-7

<p><b>Copyright information:</b></p><p>Taken from "Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse"</p><p>http://www.virologyj.com/content/4/1/118</p><p>Virology Journal 2007;4():118-118.</p><p>Published online 31 Oct 2007</p><p>PMCID:PMC2186315.</p><p></p>djacent cysteins. S1 and S2 are helper T cell peptides and M2e the 24 N-terminal aa of M2, linked through their C-terminal aa to the indicated lysines of the scaffold peptides

Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse-0

<p><b>Copyright information:</b></p><p>Taken from "Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse"</p><p>http://www.virologyj.com/content/4/1/118</p><p>Virology Journal 2007;4():118-118.</p><p>Published online 31 Oct 2007</p><p>PMCID:PMC2186315.</p><p></p>djacent cysteins. S1 and S2 are helper T cell peptides and M2e the 24 N-terminal aa of M2, linked through their C-terminal aa to the indicated lysines of the scaffold peptides

Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse-4

<p><b>Copyright information:</b></p><p>Taken from "Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse"</p><p>http://www.virologyj.com/content/4/1/118</p><p>Virology Journal 2007;4():118-118.</p><p>Published online 31 Oct 2007</p><p>PMCID:PMC2186315.</p><p></p>en squares) received PBS i.p. One day later, mice were exposed to a total respiratory tract challenge with PR8 (4 LDin 50 μl) and monitored for weight loss. Pooled data from two independent experiments are shown, each performed with 4–5 mice/group. A. Symbols show mean % body weight and SEM (relative to day 0) of 9–10 mice/group. Differences between treatment groups were tested for statistical significance at individual days. Mice treated with G2a showed significantly (p < 0,05, ANOVA) less weight loss than those treated with G1 or G2b at days 6 to 13 p.i. B. Survival. Death was defined as >30% weight loss, at which stage mice were euthanized. Differences between survival curves were tested for statistical significance by log rank test

Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse-6

<p><b>Copyright information:</b></p><p>Taken from "Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse"</p><p>http://www.virologyj.com/content/4/1/118</p><p>Virology Journal 2007;4():118-118.</p><p>Published online 31 Oct 2007</p><p>PMCID:PMC2186315.</p><p></p> μg), ODN (3 μg), CT (0.5 μg), Vir (150–200 TCIDof PR8 for primary and of Seq14 for secondary immunization), Vir(uv) (5 μg of purified uv-inactivated PR8

Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse-5

<p><b>Copyright information:</b></p><p>Taken from "Roles of adjuvant and route of vaccination in antibody response and protection engendered by a synthetic matrix protein 2-based influenza A virus vaccine in the mouse"</p><p>http://www.virologyj.com/content/4/1/118</p><p>Virology Journal 2007;4():118-118.</p><p>Published online 31 Oct 2007</p><p>PMCID:PMC2186315.</p><p></p>ction (5 μl X31, 1000 TCID). Five days after challenge, virus titers were determined in nose, trachea and lung of individual mice and the group average was determined. The average reduction in virus titer on logbasis compared to the control group (immunized with adjuvant alone) was taken as measure of strength of protection (y axis). A. Protection in nose (squares), trachea (triangles) and lung (circles) from nine groups of mice immunized by parenteral route is plotted against the M2e(pep-nat)-specific serum Ab titer (x axis). Non-linear regression analysis yielded sigmoidal regression curves with Rof 0.79 for nose (stipulated) and lung (continuous) and of 0.65 for trachea (dashed). B. Serum Ab titers and protection in nose observed in mice immunized by the i.n. route (filled squares) are plotted together with the regression line and corresponding data points (open squares) from mice immunized by parenteral route (as in A). C. Serum Ab titers and protection in lung of mice immunized by i.n. route (filled circles) are plotted together with the regression line and corresponding data points (open circles) from mice after parenteral immunization (as in A)

Immunization with M2e-MAP K2 protects against challenge with M2e-variant viruses.

<p>(<b>A</b>) Amino acid sequence comparison of M2e from influenza A/PR8 (P10) or M2e-escape mutant viruses (P10H and P10L) as described in Ref. 9. (<b>B</b>) Sera from BALB/c mice immunized i.n. three times with K2 peptide and adjuvants were assayed in an ELISA on peptides with M2e sequences as shown in (<b>A</b>) or control peptide cysBB: cysteine back-bone. (<b>C</b>) Pooled sera (1∶250 dilution) from BALB/c mice (n = 5 mice) immunized as in (<b>B</b>) were assayed in an ELISA for binding to MDCK cells infected with influenza virus A/PR/8, the M2e-escape mutants P10H and P10L, A/FM and B/Lee. The monoclonal anti-M2e Ab 14C2 (1 µg/mL) is shown as control. (<b>D</b>) BALB/c mice (n = 5 mice/group) immunized i.n. with K2 peptide and adjuvants were challenged with 1000 TCID₅₀/50 µL influenza virus A/PR/8, P10H and P10L three weeks after the third vaccine administration. Infectious virus in the lungs was determined 5 days after challenge.</p

Viral titers in lungs and nose of X31-challenged mice following vaccination.

<p>BALB/c (n = 21), C3H (n = 5), C57BL/6 (n = 3), CD1/ICR (n = 7) and Swiss Webster (n = 5) mice were immunized three times i.n. with adjuvants CpG 1826 and CT alone or K2 and adjuvants and tested for their protection against viral challenge with X31 virus (1000 TCID₅₀ in 5 µL). Virus titers in lungs and noses were determined 5 days after challenge. Results are displayed as mean ± SD.</p