Characterization of the antibody response against Plasmodium falciparum erythrocyte membrane protein 1 in human volunteers

Abstract

The immune response against the Plasmodium falciparum variant surface antigen P. falciparum erythrocyte membrane protein 1 (PfEMP1) is a key component of clinical immunity against falciparum malaria. In this study, we used sera from human volunteers who had been infected with the P. falciparum 3D7 strain to investigate the development, specificity, and dynamics of anti-PfEMP1 antibodies measured against six different strain 3D7 Duffy binding-like domain 1alpha (DBL1alpha) fusion proteins. We observed that a parasitemia of 20 to 200 infected erythrocytes per mul was required to trigger an antibody response to DBL1alpha and that antibodies against one DBL1alpha variant cross-react with other DBL1alpha variants. Both serum and purified immunoglobulin Gs (IgGs) were able to agglutinate infected erythrocytes, and purified anti-DBL1alpha IgGs bound to the live infected red blood cell surface in a punctate surface pattern, confirming that the IgGs recognize native PfEMP1. Analysis of sera from tourists naturally infected with P. falciparum suggests that the anti-PfEMP1 antibodies often persisted for more than 100 days after a single infection. These results help to further our understanding of the development of acquired immunity to P. falciparum infections.

FIG. 1.

IgG antibody reactivity to PF11_0007DBL1α in sera from volunteers B1 and B2 collected prior to (Pre) and at the indicated number of days after the first and second infections (Infn.), excluding the liver stages. Con1 and Con2 represent two different pools of Red Cross sera used as negative controls. Hyper, hyperimmune sera used as a positive control. His, anti-His antibody used to reveal protein loadings. A molecular size marker is shown on the right.

FIG. 2.

Reactivities of IgGs, purified on a PF11_0007 affinity column, to DBL1α fusion proteins. (A) Human IgG was detected in the following fractions by Western blotting using anti-human IgG: U, unbound IgG; W, wash; pH 4.5 elution; and pH 2.5 elution (lanes 1 to 8) from the PF11_0007DBL1α column. A molecular size marker is shown on the right. (B) Reactivity of serially diluted IgG in fraction 5 of the pH 2.5 elution to fusion proteins PF11_0007DBL1α, PF10_0406DBL1α, PF13_0003DBL1α, and PF07_0051DBL1α. α-His, anti-His antibody.

FIG. 3.

Serum from volunteer B2 and purified IgGs recognize full-length PfEMP1 on Western blot of parasite extract and on the live IRBC surface. (A) A Western blot of 3D7B2-IRBCs (day 23 in culture) was probed with preimmune serum from volunteer B2 (Pre-B2), postinfection serum from volunteer B2 (B2), or IgGs purified on the PF11_0007DBL1α column. Anti-ATS antibody was used as a positive control for PfEMP1, as it is a rabbit antibody against the cytoplasmic C-terminal ATS region of PfEMP1. For each blot, lane 1 represents 3D7B2 extract and lane 2 represents extract from the same number of uninfected RBCs. Molecular size markers are shown on the right. (B) Two images of live, intact 3D7-IRBCs surface labeled with IgG from volunteer B2 purified on the PF11_0007DBL1α column. The three panels show a differential interference contrast image, an immunofluorescence image, and the merged image. The arrows point to Plasmodium falciparum-infected RBCs.