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Early neutralizing IgG response to Chikungunya virus in infected patients targets a dominant linear epitope on the E2 glycoprotein.

Kam YW, Lum FM, Teo TH, Lee WW, Simarmata D, Harjanto S, Chua CL, Chan YF, Wee JK, Chow A, Lin RT, Leo YS, Le Grand R, Sam IC, Tong JC, Roques P, Wiesmüller KH, Rénia L, Rötzschke O, Ng LF - EMBO Mol Med (2012)

Bottom Line: E2EP3 is located at the N-terminus of the E2 glycoprotein and prominently exposed on the viral envelope.Screening of E2EP3 across different patient cohorts and in non-human primates demonstrated the value of this epitope as a good serology detection marker for CHIKV infection already at an early stage.Mice vaccinated by E2EP3 peptides were protected against CHIKV with reduced viremia and joint inflammation, providing a pre-clinical basis for the design of effective vaccine against arthralgia-inducing CHIKV and other alphaviruses.

View Article: PubMed Central - PubMed

Affiliation: Singapore Immunology Network, Agency for Science, Technology and Research (A*STAR), Biopolis, Singapore.

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Epitope mapping of the E2 glycoproteinCHIKV-infected patient plasma pools (Median 10 days pio) were subjected to peptide-based ELISA at a dilution of 1:2000, followed by secondary human anti-IgG-HRP using pooled peptides (P1–P11).The same set of patient plasma pools were subjected to peptide-based ELISA at a dilution of 1:2000, followed by secondary human anti-IgG-HRP using both selected peptide pools (P1, 2, 10 and 11) and individual peptides.Selected individual peptides were re-screened with patients' plasma pools at a dilution of 1:200, followed by secondary human anti-IgG3-HRP. Black solid line represents the mean value of the healthy donors and dotted line represents the value of mean ± 6 SD. Values above mean ± 6 SD are considered positive. Results represent an average of two independent experiments.
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fig02: Epitope mapping of the E2 glycoproteinCHIKV-infected patient plasma pools (Median 10 days pio) were subjected to peptide-based ELISA at a dilution of 1:2000, followed by secondary human anti-IgG-HRP using pooled peptides (P1–P11).The same set of patient plasma pools were subjected to peptide-based ELISA at a dilution of 1:2000, followed by secondary human anti-IgG-HRP using both selected peptide pools (P1, 2, 10 and 11) and individual peptides.Selected individual peptides were re-screened with patients' plasma pools at a dilution of 1:200, followed by secondary human anti-IgG3-HRP. Black solid line represents the mean value of the healthy donors and dotted line represents the value of mean ± 6 SD. Values above mean ± 6 SD are considered positive. Results represent an average of two independent experiments.

Mentions: In order to identify linear epitopes within the E2 glycoprotein, a peptide library consisting of overlapping peptides was scanned with the pooled patients' plasma (Fig 2A). The library covered the entire E2 glycoprotein and consisted of 18-mer peptides, each with an overlap of 10 amino acids. Analysis of pools combining 5 consecutive peptides revealed that the IgG-response was most pronounced against the N′-terminal part of the E2 glycoprotein (Pool P1). Only some minor reactivity was detected to the other regions of the protein (Pools P2, P10 and P11) (Fig 2A). Plasma samples were next assayed with the complete set of single peptides from each of the four active pools (Fig 2B). We found that the antibodies strongly recognized the first two peptides of pool 1. In a previous study, we established that the early IgG response against CHIKV is almost exclusively driven by antibodies of the IgG3 isotype (Kam et al, 2012). A very similar picture therefore emerged when anti-IgG3 instead of anti-IgG was used for detection (Fig 2C, Supporting information Fig 1B). Although the sensitivity of the IgG3 assay is generally weaker, the two peptides of pool 1 (P1-1 and P1-2) were clearly detectable, showing a slightly stronger titer for P1-1.


Early neutralizing IgG response to Chikungunya virus in infected patients targets a dominant linear epitope on the E2 glycoprotein.

Kam YW, Lum FM, Teo TH, Lee WW, Simarmata D, Harjanto S, Chua CL, Chan YF, Wee JK, Chow A, Lin RT, Leo YS, Le Grand R, Sam IC, Tong JC, Roques P, Wiesmüller KH, Rénia L, Rötzschke O, Ng LF - EMBO Mol Med (2012)

Epitope mapping of the E2 glycoproteinCHIKV-infected patient plasma pools (Median 10 days pio) were subjected to peptide-based ELISA at a dilution of 1:2000, followed by secondary human anti-IgG-HRP using pooled peptides (P1–P11).The same set of patient plasma pools were subjected to peptide-based ELISA at a dilution of 1:2000, followed by secondary human anti-IgG-HRP using both selected peptide pools (P1, 2, 10 and 11) and individual peptides.Selected individual peptides were re-screened with patients' plasma pools at a dilution of 1:200, followed by secondary human anti-IgG3-HRP. Black solid line represents the mean value of the healthy donors and dotted line represents the value of mean ± 6 SD. Values above mean ± 6 SD are considered positive. Results represent an average of two independent experiments.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3376860&req=5

fig02: Epitope mapping of the E2 glycoproteinCHIKV-infected patient plasma pools (Median 10 days pio) were subjected to peptide-based ELISA at a dilution of 1:2000, followed by secondary human anti-IgG-HRP using pooled peptides (P1–P11).The same set of patient plasma pools were subjected to peptide-based ELISA at a dilution of 1:2000, followed by secondary human anti-IgG-HRP using both selected peptide pools (P1, 2, 10 and 11) and individual peptides.Selected individual peptides were re-screened with patients' plasma pools at a dilution of 1:200, followed by secondary human anti-IgG3-HRP. Black solid line represents the mean value of the healthy donors and dotted line represents the value of mean ± 6 SD. Values above mean ± 6 SD are considered positive. Results represent an average of two independent experiments.
Mentions: In order to identify linear epitopes within the E2 glycoprotein, a peptide library consisting of overlapping peptides was scanned with the pooled patients' plasma (Fig 2A). The library covered the entire E2 glycoprotein and consisted of 18-mer peptides, each with an overlap of 10 amino acids. Analysis of pools combining 5 consecutive peptides revealed that the IgG-response was most pronounced against the N′-terminal part of the E2 glycoprotein (Pool P1). Only some minor reactivity was detected to the other regions of the protein (Pools P2, P10 and P11) (Fig 2A). Plasma samples were next assayed with the complete set of single peptides from each of the four active pools (Fig 2B). We found that the antibodies strongly recognized the first two peptides of pool 1. In a previous study, we established that the early IgG response against CHIKV is almost exclusively driven by antibodies of the IgG3 isotype (Kam et al, 2012). A very similar picture therefore emerged when anti-IgG3 instead of anti-IgG was used for detection (Fig 2C, Supporting information Fig 1B). Although the sensitivity of the IgG3 assay is generally weaker, the two peptides of pool 1 (P1-1 and P1-2) were clearly detectable, showing a slightly stronger titer for P1-1.

Bottom Line: E2EP3 is located at the N-terminus of the E2 glycoprotein and prominently exposed on the viral envelope.Screening of E2EP3 across different patient cohorts and in non-human primates demonstrated the value of this epitope as a good serology detection marker for CHIKV infection already at an early stage.Mice vaccinated by E2EP3 peptides were protected against CHIKV with reduced viremia and joint inflammation, providing a pre-clinical basis for the design of effective vaccine against arthralgia-inducing CHIKV and other alphaviruses.

View Article: PubMed Central - PubMed

Affiliation: Singapore Immunology Network, Agency for Science, Technology and Research (A*STAR), Biopolis, Singapore.

Show MeSH
Related in: MedlinePlus