Abstract
BackgroundProgress towards the development of a malaria vaccine against Plasmodium vivax, the most widely distributed human malaria parasite, will require a better understanding of the immune responses that confer clinical protection to patients in regions where malaria is endemic.MethodsGlutathione S-transferase (GST) and GST-fusion proteins representing the N- terminus of the merozoite surface protein 1 of P. vivax, PvMSP1-N, and the C-terminus, PvMSP1-C, were covalently coupled to BioPlex carboxylated beads. Recombinant proteins and coupled beads were used, respectively, in ELISA and Bioplex assays using immune sera of P. vivax patients from Brazil and PNG to determine IgG and subclass responses. Concordances between the two methods in the seropositivity responses were evaluated using the Kappa statistic and the Spearman's rank correlation.ResultsThe results using this methodology were compared with the classical microtitre enzyme-linked immnosorbent assay (ELISA), showing that the assay was sensitive, reproducible and had good concordance with ELISA; yet, further research into different statistical analyses seems desirable before claiming conclusive results exclusively based on multiplex assays. As expected, results demonstrated that PvMSP1 was immunogenic in natural infections of patients from different endemic regions of Brazil and Papua New Guinea (PNG), and that age correlated only with antibodies against the C-terminus part of the molecule. Furthermore, the IgG subclass profiles were different in these endemic regions having IgG3 predominantly recognizing PvMSP1 in Brazil and IgG1 predominantly recognizing PvMSP1 in PNG.ConclusionsThis study validates the use of the multiplex assay to measure naturally-acquired IgG antibodies against the merozoite surface protein 1 of P. vivax.
Highlights
Progress towards the development of a malaria vaccine against Plasmodium vivax, the most widely distributed human malaria parasite, will require a better understanding of the immune responses that confer clinical protection to patients in regions where malaria is endemic
Concordance between the BioPlex assay and enzyme-linked immnosorbent assay (ELISA) Three recombinant proteins corresponding to Glutathione S-transferase (GST) and GST-fused to the N- and C-termini of PvMSP1 were used for this analysis
Plasma samples from 87 adults from Brazil as well as from 100 children from Papua New Guinea (PNG) were tested at 1:100 dilutions to measure total IgG antibodies
Summary
Progress towards the development of a malaria vaccine against Plasmodium vivax, the most widely distributed human malaria parasite, will require a better understanding of the immune responses that confer clinical protection to patients in regions where malaria is endemic. Studies on naturally acquired immune responses against different P. vivax antigens, including, among others, merozoite surface antigens, Duffy-binding protein, circumsporozoite surface protein, apical was reported in children from Papua New Guinea [8] This latter study used a functional assay on naturallyacquired binding inhibitory antibodies against the Duffybinding protein region II (PvDBPII) to demonstrate the association of these antibodies with protection against P. vivax infection. Antibodies in these studies were measured using the ELISA assay [9], a technique requiring high amounts of coating-antigens and immune sera when large numbers of samples must be screened. These methodologies have been reported to measure antibodies to multiple malaria vaccine candidate antigens [11], to measure simultaneously antibody recognition of twenty eight Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) domains [12], and to develop a high throughput functional assay to study binding of ICAM1 to the 3D7 strain PfEMP1 repertoire [13]
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