Human antibodies to a Mr 155,000 Plasmodium falciparum antigen efficiently inhibit merozoite invasion.

Abstract

IgG from a donor clinically immune to Plasmodiumfalciparum malaria strongly inhibited reinvasion in vitro of human erythrocytes by the parasite. When added to monolayers of glutaraldehyde-fixed and air-dried erythrocytes infected with the parasite, this IgG also displayed a characteristic immunofluorescence restricted to the surface of infected erythrocytes. Elution of the IgG adsorbed to such monolayers gave an antibody fraction that was 40 times more efficient in the reinvasion inhibition assay (50% inhibition titer, <1 ,ug/ml) than the original IgG preparation. falciparum (1, 2). These antigens are released from bursting schizonts or merozoites and are deposited in the RBC membrane in the course of invasion. Antibodies recognizing these antigens are of high prevalence in sera from residents of a holoendemic area of Africa (Liberia) but are also present in many sera of acutely infected patients from different parts of the world. A polypeptide of Mr 155,000 (Pf 155) appears to be primarily responsible for this RBC membrane IF. A similar antigen of the same apparent molecular weight has also been found recently by two other groups (3, 4). To further investigate the function of these antigens and the possible significance of the immune response against them, we have now tested the antibodies reacting with Pf 155 and related minor components for their capacity to inhibit RBC invasion by merozoites in vitro. MATERIAL AND METHODSParasites. Parasites were from a Tanzanian strain of P. falciparum (F 32) isolated in 1978 (5) and cultured in vitro in blood group O RBCs (6).Immune Sera. Five sera (Kinon, IS-6 to -8, X-12) were from Liberians, >15 years old (except X-12, from a 12-yearold boy), living in a P. falciparum holoendemic area in which in adult life clinical illness is rare and parasitemia is kept at a low-grade, mostly subpatent, level despite high sporozoite inoculation rates (7). These donors, designated as clinically immune, had not taken any antimalarial drug for the last 6 weeks (X-12) or longer. The 6 remaining donors had acute P. falciparum infections. Four donors (ASF, MV, JOR, FG) were South Americans from an endemic area of Colombia. Two Swedish patients (YC, HP) were suffering from a first infection acquired in Kenya. IgG was prepared by ammonium sulfate precipitation and ion-exchange chromatography on DEAE-Sephadex (Pharmacia). Immunoglobulin concentrations were determined by ELISA.In Vitro Growth Inhibition Assay. Infected RBCs from P. falciparum cultures (5-10% parasitemia, -70% schizonts) were diluted with normal O RBCs to a parasitemia of 0.5%. They were adjusted to 2% hematocrit with Hepes-buffered (20 mM) RPMI-1640 medium (GIBCO) containing 15% normal human serum, 2 mM glutamine, 25 pg of gentamycin per ml, and 0.2% NaHCO3 [complete tissue culture medium (TC medium)]. In some experiments, synchronized parasite cultures were used. In this case, the original cultures (5-10% parasitemia) were adjusted to 10% hematocrit and layered on top of 2.5 ml of 60% Percoll (Pharmacia) diluted in com...

Antibacterial and antimalarial properties of peptides that are cecropin‐melittin hybrids

“…Inhibition of the growth of the blood stream forms of P. falciparium by different amounts of hybrid 2, CA(l-13)M(l-13), and four other peptides used for comparison. Details of the assay are given in section 2 and in [19]. Duplicate assays were run for hybrid 2.…”

“…The blood stream forms of the malaria parasite, Plasmodium falciparium, primarily late trophozoites and early schizonts, were assayed by recording the inhibition of the reinvasion of human red cells [19]. Quadruplicate microcultures of strain F32 (Tanzania) were incubated for 20 h at 37°C in a complete tissue culture medium, with or without different concentrations of the peptides to be assayed.…”

Synthetic gene construct expressing a repeated and highly immunogenic epitope of the Plasmodium falciparum antigen Pf155.

“…Recently, we have identified a P. falciparum antigen of Mr 155,000 (designated Pf155) that is deposited in the erythrocyte (RBC) membrane during merozoite invasion (6,7). Human antibodies to this merozoite-associated antigen were shown to efficiently inhibit P. falciparum reinvasion in in vitro cultures (8). In addition, a correlation was found between high levels of antibodies to Pf155 and acquired P. falciparum immunity in children living in a holoendemic area of Africa (9).…”

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

“…Cultures at a ®nal parasitaemia of 1% and a haematocrit of 2% were set up in microculture plates. The cultures were mixed with malaria culture medium or supplemented with serial dilutions of different af®nity-puri®ed antibodies and were incubated at 378C for 20 h by the candle jar method [36]. W150 42 39 73 54 30 95 85 75 10 W101 27 26 94 35 20 87 75 73 15 bd191 34 25 87 28 21 92 75 79 12 T42 50 69 84 41 28 97 69 70 15 ts46 33 28 92 19 17 90 70 86 15 * Peptides used for serum absorption.…”