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Multi-Dimensional Measurement of Antibody-Mediated Heterosubtypic Immunity to Influenza.

Wang J, Hilchey SP, Hyrien O, Huertas N, Perry S, Ramanunninair M, Bucher D, Zand MS - PLoS ONE (2015)

Bottom Line: Pre- and post-intramuscular trivalent influenza vaccination levels of virus specific Ig were consistent with HAI titer and ELISA measurements.Results were in good agreement with a phylogenetic tree generated from hierarchical clustering of the genomic HA sequences.This is the first report of the use of a multiplex method for antigenic cartography using ferret sera.

View Article: PubMed Central - PubMed

Affiliation: Division of Nephrology, Department of Medicine and the Rochester Center for Biodefense Immune Modeling, University of Rochester Medical Center, Rochester, New York, United States of America.

ABSTRACT
The human immune response to influenza vaccination depends in part on preexisting cross-reactive (heterosubtypic) immunity from previous infection by, and/or vaccination with, influenza strains that share antigenic determinants with the vaccine strains. However, current methods for assessing heterosubtypic antibody responses against influenza, including the hemagglutination-inhibition (HAI) assay and ELISA, are time and labor intensive, and require moderate amounts of serum and reagents. To address these issues we have developed a fluorescent multiplex assay, mPlex-Flu, that rapidly and simultaneously measures strain specific IgG, IgA, and IgM antibodies against influenza hemagglutinin (HA) from multiple viral strains. We cloned, expressed and purified HA proteins from 12 influenza strains, and coupled them to multiplex beads. Assay validation showed that minimal sample volumes (<5 μl of serum) were needed, and the assay had a linear response over a four Log10 range. The assay detected nanogram levels of anti-influenza specific antibodies, had high accuracy and reproducibility, with an average percentage coefficient of variation (%CV) of 9.06 for intra-assay and 12.94 for inter-assay variability. Pre- and post-intramuscular trivalent influenza vaccination levels of virus specific Ig were consistent with HAI titer and ELISA measurements. A significant advantage of the mPLEX-Flu assay over the HAI assay is the ability to perform antigenic cartography, determining the antigenic distances between influenza HA's, without mathematical correction for HAI data issues. For validation we performed antigenic cartography on 14 different post-influenza infection ferret sera assayed against 12 different influenza HA's. Results were in good agreement with a phylogenetic tree generated from hierarchical clustering of the genomic HA sequences. This is the first report of the use of a multiplex method for antigenic cartography using ferret sera. Overall, the mPlex-Flu assay provides a powerful tool to rapidly assess the influenza antibody repertoire in large populations and to study heterosubtypic immunity induced by influenza vaccination.

No MeSH data available.


Related in: MedlinePlus

The binding reaction (MFI) of the mPlex-Flu assay using influenza ferret virus anti-serum and anti-influenza monoclonal antibodies.A panel of 12 rHA coupled-beads: A/California/07/2009 (A/Cal09) and A/South Carolina/1/1918 (A/S Car1918) H1 substrains, A/Perth/16/2009 (A/Perth09), A/Victoria/210/2009 (A/Vic09), A/Victoria/361/2011 (A/Vic11), A/Texas/50/2012 (A/Tex13), and A/Hiroshima/2005 (A/Hir05) A/Hong Kong/01/1968 (A/HK68) and A/Port Chalmers/1/1973 (A/P.C73) H3 substrains, B/Brisbane/60/2008 (B/Bri08), B/Wisconsin/1/2010 (B/Wis10), B/Massachusetts/2/2012 (B/Mass12) B strains, was used to measure antibody levels from; (a) post-infectious ferret antisera of influenza vaccine associated strains obtained from the Influenza Reagent Resource (IRR). Shown are the MFIs generated from the twelve-bead set assay utilizing dilutions based on the HAI titers provided by IRR. (b) The MFIs generated using mouse monoclonal antibodies directed against specific rHAs or whole influenza provided by the IRR. The concentration of all monoclonal antibodies in the assays is 2.5 μg/ml (1:200 dilution of 500 μg/ml). The MFI represents the mean of triplicates measurements.
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pone.0129858.g005: The binding reaction (MFI) of the mPlex-Flu assay using influenza ferret virus anti-serum and anti-influenza monoclonal antibodies.A panel of 12 rHA coupled-beads: A/California/07/2009 (A/Cal09) and A/South Carolina/1/1918 (A/S Car1918) H1 substrains, A/Perth/16/2009 (A/Perth09), A/Victoria/210/2009 (A/Vic09), A/Victoria/361/2011 (A/Vic11), A/Texas/50/2012 (A/Tex13), and A/Hiroshima/2005 (A/Hir05) A/Hong Kong/01/1968 (A/HK68) and A/Port Chalmers/1/1973 (A/P.C73) H3 substrains, B/Brisbane/60/2008 (B/Bri08), B/Wisconsin/1/2010 (B/Wis10), B/Massachusetts/2/2012 (B/Mass12) B strains, was used to measure antibody levels from; (a) post-infectious ferret antisera of influenza vaccine associated strains obtained from the Influenza Reagent Resource (IRR). Shown are the MFIs generated from the twelve-bead set assay utilizing dilutions based on the HAI titers provided by IRR. (b) The MFIs generated using mouse monoclonal antibodies directed against specific rHAs or whole influenza provided by the IRR. The concentration of all monoclonal antibodies in the assays is 2.5 μg/ml (1:200 dilution of 500 μg/ml). The MFI represents the mean of triplicates measurements.

Mentions: We also explored the ability of the mPlex-flu assay to rapidly characterize the antigenic similarity between influenza HAs from disparate viral strains by antigenic cartography. Given that the mPlex-Flu assay can simultaneously measure reactivity against multiple influenza strains with a continuous quantitative measure, high sensitivity, and a 4 log10 range, we were interested to know how it would perform for rapid assessment of antigenic similarity among multiple disparate influenza strains. We tested a panel of ferret anti-influenza sera (Table 2) and murine monoclonal antibodies derived from single strain influenza rHA vaccinations (Table 3) against 12 rHA using the mPlex-Flu assay. This included rHAs from all eight influenza vaccine strains covered by the 2010–2014 TIV vaccines (A/California/07/2009, A/Perth/16/2009, A/Victoria/210/2009, A/Victoria/361/2011, A/Texas/50/2012, B/Brisbane/60/2008, B/Wisconsin/1/2010, B/Massachusetts/2/2012), three more temporally distant H3 control strains (A/Port Chalmers /1973, A/Hong Kong/1968 and A/Hiroshima/2005), and one temporally distant H1 control strain (A/South Carolina/1/1918). The results are shown in Fig 5. We found that A/California/07/09 rHA was strongly bound by homologous post-influenza vaccine ferret antisera (FR-359) but only weakly by H1 subtype (H1 strain) anti-A/Brisbane/59/2007 sera (FR-388 and FR-288), reflecting the antigenic shift of this pandemic virus. In contrast, the historical H1 control strain A/South Carolina/1/1918 showed stronger binding by the anti-A/Brisbane/59/2007 sera than anti-A/California/07/2009 post-infectious, suggesting that the antigenicity of A/South Carolina/1/1918 is closer to A/Brisbane/59/2007 than A/California/07/2009, at least in the context of primary influenza infection.


Multi-Dimensional Measurement of Antibody-Mediated Heterosubtypic Immunity to Influenza.

Wang J, Hilchey SP, Hyrien O, Huertas N, Perry S, Ramanunninair M, Bucher D, Zand MS - PLoS ONE (2015)

The binding reaction (MFI) of the mPlex-Flu assay using influenza ferret virus anti-serum and anti-influenza monoclonal antibodies.A panel of 12 rHA coupled-beads: A/California/07/2009 (A/Cal09) and A/South Carolina/1/1918 (A/S Car1918) H1 substrains, A/Perth/16/2009 (A/Perth09), A/Victoria/210/2009 (A/Vic09), A/Victoria/361/2011 (A/Vic11), A/Texas/50/2012 (A/Tex13), and A/Hiroshima/2005 (A/Hir05) A/Hong Kong/01/1968 (A/HK68) and A/Port Chalmers/1/1973 (A/P.C73) H3 substrains, B/Brisbane/60/2008 (B/Bri08), B/Wisconsin/1/2010 (B/Wis10), B/Massachusetts/2/2012 (B/Mass12) B strains, was used to measure antibody levels from; (a) post-infectious ferret antisera of influenza vaccine associated strains obtained from the Influenza Reagent Resource (IRR). Shown are the MFIs generated from the twelve-bead set assay utilizing dilutions based on the HAI titers provided by IRR. (b) The MFIs generated using mouse monoclonal antibodies directed against specific rHAs or whole influenza provided by the IRR. The concentration of all monoclonal antibodies in the assays is 2.5 μg/ml (1:200 dilution of 500 μg/ml). The MFI represents the mean of triplicates measurements.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4478018&req=5

pone.0129858.g005: The binding reaction (MFI) of the mPlex-Flu assay using influenza ferret virus anti-serum and anti-influenza monoclonal antibodies.A panel of 12 rHA coupled-beads: A/California/07/2009 (A/Cal09) and A/South Carolina/1/1918 (A/S Car1918) H1 substrains, A/Perth/16/2009 (A/Perth09), A/Victoria/210/2009 (A/Vic09), A/Victoria/361/2011 (A/Vic11), A/Texas/50/2012 (A/Tex13), and A/Hiroshima/2005 (A/Hir05) A/Hong Kong/01/1968 (A/HK68) and A/Port Chalmers/1/1973 (A/P.C73) H3 substrains, B/Brisbane/60/2008 (B/Bri08), B/Wisconsin/1/2010 (B/Wis10), B/Massachusetts/2/2012 (B/Mass12) B strains, was used to measure antibody levels from; (a) post-infectious ferret antisera of influenza vaccine associated strains obtained from the Influenza Reagent Resource (IRR). Shown are the MFIs generated from the twelve-bead set assay utilizing dilutions based on the HAI titers provided by IRR. (b) The MFIs generated using mouse monoclonal antibodies directed against specific rHAs or whole influenza provided by the IRR. The concentration of all monoclonal antibodies in the assays is 2.5 μg/ml (1:200 dilution of 500 μg/ml). The MFI represents the mean of triplicates measurements.
Mentions: We also explored the ability of the mPlex-flu assay to rapidly characterize the antigenic similarity between influenza HAs from disparate viral strains by antigenic cartography. Given that the mPlex-Flu assay can simultaneously measure reactivity against multiple influenza strains with a continuous quantitative measure, high sensitivity, and a 4 log10 range, we were interested to know how it would perform for rapid assessment of antigenic similarity among multiple disparate influenza strains. We tested a panel of ferret anti-influenza sera (Table 2) and murine monoclonal antibodies derived from single strain influenza rHA vaccinations (Table 3) against 12 rHA using the mPlex-Flu assay. This included rHAs from all eight influenza vaccine strains covered by the 2010–2014 TIV vaccines (A/California/07/2009, A/Perth/16/2009, A/Victoria/210/2009, A/Victoria/361/2011, A/Texas/50/2012, B/Brisbane/60/2008, B/Wisconsin/1/2010, B/Massachusetts/2/2012), three more temporally distant H3 control strains (A/Port Chalmers /1973, A/Hong Kong/1968 and A/Hiroshima/2005), and one temporally distant H1 control strain (A/South Carolina/1/1918). The results are shown in Fig 5. We found that A/California/07/09 rHA was strongly bound by homologous post-influenza vaccine ferret antisera (FR-359) but only weakly by H1 subtype (H1 strain) anti-A/Brisbane/59/2007 sera (FR-388 and FR-288), reflecting the antigenic shift of this pandemic virus. In contrast, the historical H1 control strain A/South Carolina/1/1918 showed stronger binding by the anti-A/Brisbane/59/2007 sera than anti-A/California/07/2009 post-infectious, suggesting that the antigenicity of A/South Carolina/1/1918 is closer to A/Brisbane/59/2007 than A/California/07/2009, at least in the context of primary influenza infection.

Bottom Line: Pre- and post-intramuscular trivalent influenza vaccination levels of virus specific Ig were consistent with HAI titer and ELISA measurements.Results were in good agreement with a phylogenetic tree generated from hierarchical clustering of the genomic HA sequences.This is the first report of the use of a multiplex method for antigenic cartography using ferret sera.

View Article: PubMed Central - PubMed

Affiliation: Division of Nephrology, Department of Medicine and the Rochester Center for Biodefense Immune Modeling, University of Rochester Medical Center, Rochester, New York, United States of America.

ABSTRACT
The human immune response to influenza vaccination depends in part on preexisting cross-reactive (heterosubtypic) immunity from previous infection by, and/or vaccination with, influenza strains that share antigenic determinants with the vaccine strains. However, current methods for assessing heterosubtypic antibody responses against influenza, including the hemagglutination-inhibition (HAI) assay and ELISA, are time and labor intensive, and require moderate amounts of serum and reagents. To address these issues we have developed a fluorescent multiplex assay, mPlex-Flu, that rapidly and simultaneously measures strain specific IgG, IgA, and IgM antibodies against influenza hemagglutinin (HA) from multiple viral strains. We cloned, expressed and purified HA proteins from 12 influenza strains, and coupled them to multiplex beads. Assay validation showed that minimal sample volumes (<5 μl of serum) were needed, and the assay had a linear response over a four Log10 range. The assay detected nanogram levels of anti-influenza specific antibodies, had high accuracy and reproducibility, with an average percentage coefficient of variation (%CV) of 9.06 for intra-assay and 12.94 for inter-assay variability. Pre- and post-intramuscular trivalent influenza vaccination levels of virus specific Ig were consistent with HAI titer and ELISA measurements. A significant advantage of the mPLEX-Flu assay over the HAI assay is the ability to perform antigenic cartography, determining the antigenic distances between influenza HA's, without mathematical correction for HAI data issues. For validation we performed antigenic cartography on 14 different post-influenza infection ferret sera assayed against 12 different influenza HA's. Results were in good agreement with a phylogenetic tree generated from hierarchical clustering of the genomic HA sequences. This is the first report of the use of a multiplex method for antigenic cartography using ferret sera. Overall, the mPlex-Flu assay provides a powerful tool to rapidly assess the influenza antibody repertoire in large populations and to study heterosubtypic immunity induced by influenza vaccination.

No MeSH data available.


Related in: MedlinePlus