Antibodies in malarial sera to parasite antigens in the membrane of erythrocytes infected with early asexual stages of Plasmodium falciparum.

Abstract

Monolayers of human erythrocytes (E) infected with Plasmodium falciparum were briefly fixed with 1% glutaraldehyde and air dried. They were then exposed to sera from patients with P. falciparum malaria or from donors immune to this parasite and tested in an indirect immunofluorescence assay (IFA). Parasites in infected E were made visible by counterstaining with ethidium bromide. Immunofluorescence (IF) was restricted to the surface of infected E. No antibody binding was detected unless the E were dried, suggesting that the relevant antigens were not available on the outer layers of the E surface. Staining over large parts of the E surface was seen already when the merozoite penetrated noninfected cells and was strong in E containing early stages of the parasite (rings, trophozoites). It was weak or absent from E containing schizonts. Antibodies in sera from different parts of Africa, Colombia, or Sweden reacted similarly with E infected with a Tanzanian P. falciparum strain kept in culture for many years and with parasitized E freshly drawn from African, Swedish, or Colombian patients. All sera from residents of a holoendemic area (Liberia) were IFA positive. In contrast, some sera from Colombian or Swedish patients with primary infection gave negative results. The results of the IFA and of an enzyme-linked immunosorbent assay in which fixed and dried E were the targets were well-correlated, suggesting that the same antibodies were detected by these assays. The antigens involved in the IFA were susceptible to pronase but not to trypsin or neuraminidase. E surface IF was inhibited by lysates of infected E, merozoite extracts, or soluble antigens present in P. falciparum culture supernatants but not by lysates of normal E or ghost extracts. The inhibitory antigens were heat stable (100 degrees C, 5 min). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by immunoblotting of either antigen-enriched preparations from culture supernatants or merozoite extracts showed that antibodies eluted from monolayers of infected E reacted consistently with a predominant polypeptide of Mr 155,000 and two to four minor polypeptides of lower molecular weights. Metabolic labeling of the parasites with 75Se-methionine indicated that these antigens were parasite derived. We conclude that the antigens involved in these reactions are released from bursting schizonts or merozoites and are deposited in the E membrane in the course of invasion.(ABSTRACT TRUNCATED AT 400 WORDS)

Immune response to Plasmodium falciparum antigens in Cameroonian primigravidae: evolution after delivery and during second pregnancy

“…Anti-Pf155/RESA antibodies were measured by erythrocyte membrane immunofluorescent (EMIF) assay using a glutaraldehyde-fixed and air-dried monolayer of ring-infected erythrocytes (threshold of positivity, a reciprocal titre of 4) [9]. Anti-peptide antibodies were tested by FAST-ELISA using peptides conjugated to bovine serum albumin (BSA), as described [19].…”

“…Moreover, the IL-2 in vitro production in response to Pf155/ RESA, a major P. falciparum antigen [9], was reduced. IL-4 and interferon-gamma (IFN-g) in vitro production did not appear to be affected in primigravidae [10,11].…”

Analysis of the T‐Cell Receptor Vβ Usage in Monozygotic and Dizygotic Twins Living in a Plasmodium falciparum Endemic Area in West Africa

“…Eight donors were parasitaemic (geometric means asexual parasitaemia, 31/ml), although none was clinically unwell, indicating that the donors had some degree of immunity. All donors had anti-P. falciparum antibody titres Ն 1:1000 (with 32% Ն 1:25000), reflecting heavy exposure to P. falciparum in the past [22].…”

Antibodies to a non-repeat region ofPlasmodium falciparumantigen Pf155/RESA in individuals from malaria-endemic areas

“…Indirect immuno¯uorescence assay (IFA) was performed on monolayers of either air-dried parasitized erythrocytes or airdried parasitized erythrocytes treated with glutardialdehyde (GDA) [33]. The antibodies to be tested were added in serial dilutions and incubated for 30 min.…”

“…Bound antibodies were detected by using biotinylated goat anti-human IgG and avidin conjugated with FITC (Vector Labs Inc., Burlingame, CA). Fixed slides were counterstained with ethidium bromide to visualize parasites inside the erythrocytes [33].…”