Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2011 Jun 20:10:168.
doi: 10.1186/1475-2875-10-168.

Measuring naturally acquired immune responses to candidate malaria vaccine antigens in Ghanaian adults

Daniel Dodoo  1 , Michael R Hollingdale, Dorothy Anum, Kwadwo A Koram, Ben Gyan, Bartholomew D Akanmori, Josephine Ocran, Susan Adu-Amankwah, Harini Geneshan, Esteban Abot, Jennylyn Legano, Glenna Banania, Renato Sayo, Donald Brambilla, Sanjai Kumar, Denise L Doolan, William O Rogers, Judith Epstein, Thomas L Richie, Martha Sedegah
Affiliations

Measuring naturally acquired immune responses to candidate malaria vaccine antigens in Ghanaian adults

Daniel Dodoo et al. Malar J. .

Abstract

Background: To prepare field sites for malaria vaccine trials, it is important to determine baseline antibody and T cell responses to candidate malaria vaccine antigens. Assessing T cell responses is especially challenging, given genetic restriction, low responses observed in endemic areas, their variability over time, potential suppression by parasitaemia and the intrinsic variability of the assays.

Methods: In Part A of this study, antibody titres were measured in adults from urban and rural communities in Ghana to recombinant Plasmodium falciparum CSP, SSP2/TRAP, LSA1, EXP1, MSP1, MSP3 and EBA175 by ELISA, and to sporozoites and infected erythrocytes by IFA. Positive ELISA responses were determined using two methods. T cell responses to defined CD8 or CD4 T cell epitopes from CSP, SSP2/TRAP, LSA1 and EXP1 were measured by ex vivo IFN-γ ELISpot assays using HLA-matched Class I- and DR-restricted synthetic peptides. In Part B, the reproducibility of the ELISpot assay to CSP and AMA1 was measured by repeating assays of individual samples using peptide pools and low, medium or high stringency criteria for defining positive responses, and by comparing samples collected two weeks apart.

Results: In Part A, positive antibody responses varied widely from 17%-100%, according to the antigen and statistical method, with blood stage antigens showing more frequent and higher magnitude responses. ELISA titres were higher in rural subjects, while IFA titres and the frequencies and magnitudes of ex vivo ELISpot activities were similar in both communities. DR-restricted peptides showed stronger responses than Class I-restricted peptides. In Part B, the most stringent statistical criteria gave the fewest, and the least stringent the most positive responses, with reproducibility slightly higher using the least stringent method when assays were repeated. Results varied significantly between the two-week time-points for many participants.

Conclusions: All participants were positive for at least one malaria protein by ELISA, with results dependent on the criteria for positivity. Likewise, ELISpot responses varied among participants, but were relatively reproducible by the three methods tested, especially the least stringent, when assays were repeated. However, results often differed between samples taken two weeks apart, indicating significant biological variability over short intervals.

PubMed Disclaimer

Figures

Figure 1
Figure 1
IFA titers of sera from volunteers in Accra or Mampong compared with naïve controls from the US. Horizontal bars represent geomean titers and filled circles represent individual titers.
Figure 2
Figure 2
ELISA titers of sera from volunteers in Accra or Mampong compared with naïve controls from Denmark. Horizontal bars represent geometric mean titers and filled circles represent individual titers. P values are the differences in activity for each antigen between urban and rural Ghana populations; the only significant difference was for MSP3.
Figure 3
Figure 3
The frequency distribution of the medium controls (sfc/m PBMC) for all the CSP and AMA1 assays. The numbers of medium controls from each assay are distributed against the ELISpot activity (sfc/m). The distribution of medium controls from all assays using CSP or AMA1 peptide pools, and the total using both antigens, show the same distribution.
Figure 4
Figure 4
Method 1: mean ELISpot activity of all positive assays of peptide pool stimulated PBMC compared with medium-only controls. ELISpot activities of each set of individual volunteers are plotted against their corresponding medium only controls. Each symbol represents the mean of the triplicates and the black dotted line represents >55 sfc/m activity.
Figure 5
Figure 5
Positive ELISpot activities with CSP or AMA1 peptide pools with Methods 1, 2 or 3. The top three rows for each volunteer denote the three replicate assays for the first time point, and the second three rows denote the three replicate assays for the second time-point. An assay is considered positive if at least one peptide pool was positive using that method. All positive CSP and AMA1 activities are shown in light or medium gray respectively. The responses of the five volunteers (v1024, v1036, v1039, v1054 and v1056, for whom both CSP and AMA1 assays were performed) were combined and shown in dark gray. *Assay not done.
Figure 6
Figure 6
Method 1: Linkage of ELISpot activities. Left panel: ELISpot activities of Ap7 and Ap9 show moderate linkage (r2 = 0,63). Right panel: no linkage between Ap7 and Cp1 (r2 = 0.03).
Figure 7
Figure 7
Comparison of positive assays using Methods 1, 2 and 3. The number of time-points with assays that were all positive (3/3), all negative (0/3) or 1/3 or 2/3 positive using Methods 1, 2 or 3 are expressed as percent of total number. For CSP and AMA1, the numbers of time-points that were 3/3 positive was highest using Method 1 and dropped with Method 2 and was lowest with Method 3. Conversely, the number of time-points that were all 3/3 negative was lowest with Method 1 and rose with Method 2 and was highest with Method 3. When CSP and AMA1 were combined, Method 1 identified more 3/3 positive or 3/3 negative (0/3 positive) time-points than Methods 2 or 3.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Todryk SM, Bejon P, Mwangi T, Plebanski M, Urban B, Marsh K, Hill AV, Flanagan KL. Correlation of memory T cell responses against TRAP with protection from clinical malaria, and CD4 CD25 high T cells with susceptibility in Kenyans. PLoS ONE. 2008;3:e2027. doi: 10.1371/journal.pone.0002027. - DOI - PMC - PubMed
    1. Fowkes FJ, Richards JS, Simpson JA, Beeson JG. The relationship between anti-merozoite antibodies and incidence of Plasmodium falciparum malaria: A systematic review and meta-analysis. PLoS Med. p. e1000218. - PMC - PubMed
    1. John CC, Tande AJ, Moormann AM, Sumba PO, Lanar DE, Min XM, Kazura JW. Antibodies to pre-erythrocytic Plasmodium falciparum antigens and risk of clinical malaria in Kenyan children. J Infect Dis. 2008;197:519–526. doi: 10.1086/526787. - DOI - PMC - PubMed
    1. Webster HK, Boudreau EF, Pang LW, Permpanich B, Sookto P, Wirtz RA. Development of immunity in natural Plasmodium falciparum malaria: antibodies to the falciparum sporozoite vaccine 1 antigen (R32tet32) J Clin Microbiol. 1987;25:1002–1008. - PMC - PubMed
    1. Charoenvit Y, Leef MF, Yuan LF, Sedegah M, Beaudoin RL. Characterization of Plasmodium yoelii monoclonal antibodies directed against stage-specific sporozoite antigens. Infect Immun. 1987;55:604–608. - PMC - PubMed

Publication types

MeSH terms