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H Perlmann

Publications and source records attributed to H Perlmann.

At least 55 records · Page 3Linked to original sources
PubMed1990

Anti-idiotypic antibodies counteract the invasion inhibition capacity of antibodies to major epitopes of the Plasmodium falciparum antigen Pf155/RESA.

Rabbits were immunized with the synthetic peptide EENVEHDA or (EENV)2, corresponding to a tandemly repeated sequence in the C-terminal part of the Plasmodium falciparum antigen Pf155/RESA, or with Escherichia coli-derived fusion proteins containing the corresponding repeats. For all sera, the capacity of the total immunoglobulin G fractions to inhibit P. falciparum merozoite invasion in vitro was similar and relatively low. Affinity purification of Pf155/RESA-specific antibodies on parasite-infected erythrocyte monolayers or on peptide columns increased the inhibitory capacity 50 to 5,000 times, whereas the immunofluorescence titers were increased only 10 times. The addition of small amounts of total immunoglobulin G to the affinity-purified antibodies gave a marked and dose-dependent reduction of the inhibitory capacity of the purified antibodies. However, this reduction was only seen in combinations where the immunoglobulin G fraction was from the same serum as the affinity-purified antibodies, suggesting that it was mediated by anti-idiotypic antibodies reacting with non-cross-reacting idiotopes of the invasion-inhibiting antibodies.

Amino Acid Sequence↗
PubMed1989

T and B cell responses of Plasmodium falciparum malaria-immune individuals to synthetic peptides corresponding to sequences in different regions of the P. falciparum antigen Pf155/RESA.

The C-terminal (3') amino acid repeat region of the Plasmodium falciparum Ag Pf155/RESA, a vaccine candidate, contains immunodominant T and B cell epitopes. In order to identify additional T cell epitopes in the molecule, synthetic peptides corresponding to the centrally (5') located repeat region, as well as to four nonrepeated regions, were synthesized. T cells from 46 P. falciparum-primed individuals living in a holoendemic area of The Gambia where malaria transmission is seasonal were tested for their responsiveness to the peptides by thymidine incorporation and IFN-gamma release. There was a considerable variation in the response to different peptides. Proliferation and IFN-gamma release were not correlated in individual donors, underlining the importance of measuring both activities when screening donor populations for total T cell responsiveness to a given Ag. Whereas 72% of the donors responded with proliferation and/or IFN-gamma release to the intact protein the mean % responders to the peptides was 40%. The most frequent responses (up to 60%) were induced with peptides from the 3'- and 5'-repeat region of the protein. Analysis of some closely related sequences in the 3'-repeat region indicated that they contained at least two epitopes that were either distinct or cross-reacting in different donors, suggesting difference in the genetic control of these responses. When the same peptides were investigated for reactivity with antibodies, the best T cell inducing sequences also displayed the best antibody reactivities. However, in individual donors, T and B cell responses were not correlated. T cell responses were shown to persist after a period with no P. falciparum transmission, whereas antibody concentrations tended to decrease, suggesting differences in the requirements of boosting at the T and B cell levels, respectively.

Amino Acid Sequence↗
PubMed1989

Dissection of the human antibody response to the malaria antigen Pf155/RESA into epitope specific components.

The development of vaccines is presently receiving major attention in malaria research. As it is not possible to base malaria vaccines on the use of killed or attenuated organisms, the vaccines which are being developed are subunit vaccines in which the immunogens consist of defined parasite antigens or antigenic fragments. Since protective immunity to malaria involves both antibody-dependent and antibody-independent mechanisms, the immunogens in a subunit vaccine must have the capacity to induce relevant B- and T-cell responses in the majority of vaccinees. In turn, this requires good knowledge of these responses in humans who have acquired immunity through natural infection. In this paper we have summarized our recent work on the dissection into epitope-specific components of the human antibody response to the Plasmodium falciparum antigen Pf155/RESA, a recognized candidate for a vaccine against the asexual blood stages of this parasite. Epitope mapping of the antigen by means of short synthetic peptides led to the identification in several molecular regions of short amino acid sequences constituting linear and probably immunodominant B-cell epitopes. The antigenically most active region was located in the C-terminus of the molecule. This region, which consists of approximately 40 related, 4- or 8-amino acid long repeats, induced higher antibody concentrations in a larger number of malaria-immune donors than any of the other regions. A large fraction of these antibodies bound to short synthetic peptides representing the major repeat motifs of Pf155/RESA. Although these repeats are made up of closely related amino acid sequences, the antibody response to them was highly polyclonal, indicating the presence of several linear and probably also conformational epitopes which gave rise to a variety of cross-reacting as well as monospecific antibodies. Further analysis revealed that the levels of antibodies differing in specificity and/or avidity for different peptides varied independently of each other in individual donors. In an area (Liberia) where malaria transmission is holoendemic and perennial, these antibody profiles remained constant when individual donors were followed over several years. Since the C-terminal repeat region of Pf155/RESA is conserved in different P. falciparum strains, the results reflect differences in the genetic regulation of epitope-specific host responses rather than antigenic differences between infecting parasites. In donors living in an area with high but seasonal malaria transmission, antibody levels usually drop to lower levels when there is no transmission.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1989

An epidemiological study of humoral and cell-mediated immune response to the Plasmodium falciparum antigen PF155/RESA in adult Liberians.

We investigated the seroreactivity and T cell reactivity against the Plasmodium falciparum antigen Pf155/RESA, different oligopeptides from the 3' and 5' repeat regions of the Pf155/RESA antigen, and crude Plasmodium falciparum antigens in 164 adult Liberians. We compared 2 long-term residential groups with high and low exposure to malaria. The seropositive rate to the peptides was significantly higher with increased exposure. There was no significant difference in response rates to the Pf155/RESA. This may indicate the level of persistent T cell memory in previously primed donors. The seropositive rates to 3 Pf155/RESA peptides and the rates measured by either 3H-thymidine incorporation or IFN-gamma release after stimulation with Pf155/RESA and the peptides were all lower in parasite positive individuals. Even low grade, asymptomatic parasitemia can impair the T cell response in vitro. The lower antibody response in parasite positive subjects may be explained by either antibody consumption or lower protection against malaria parasitemia in subjects with low concentrations of antibodies against the Pf155/RESA antigen.

Adult↗
PubMed1989

Phenotypic characterization of mononuclear white cells using finger-prick blood and light microscopy.

A technique for the enumeration of T cell subsets in capillary blood for use in the field is described. Mononuclear white cells were separated from 0.4 ml capillary blood, kept covered by culture medium on a Multitest slide, and incubated with monoclonal antibodies to the CD2, CD4, CD8, and interleukin-2 receptor antigens. Secondary and tertiary alkaline phosphatase conjugates were used for labeling. The substrate gave a distinct blue surface staining to the positive lymphocytes. Comparison was made with a conventional immunofluorescent antibody technique (r = 0.95).

Adult↗
PubMed1989

The seroreactivity against Pf155 (RESA) antigen in villagers from a mesoendemic area in Somalia.

Pf155 (RESA), a malaria antigen in the membrane of infected red cells, was recently identified as a possible future malaria vaccine candidate. In this study the seroreactivities against this antigen were compared with those against crude parasitic antigens in 195 subjects from a Somali village with mesoendemic malaria. The seroreactivities were determined with immunofluorescence. With age, there was an increased seroreactivity to both Pf155 and the crude parasitic antigens. However, the acquisition of seroreactivity was much slower against Pf155. Hence in the age group 15-24 years, only half of the subjects had detectable antibodies against Pf155.

Adolescent↗
PubMed1988

T cell reactivity of defined peptides from a major Plasmodium falciparum vaccine candidate: the Pf155/RESA antigen.

Several immunodominant B-cell epitopes of the P. falciparum antigen blood stage Pf155/RESA, a major vaccine candidate antigen, are located in the molecular regions containing amino acid repeats. We started to map Pf155/RESA for T cell reactive epitopes. For this purpose, short synthetic peptides corresponding to the 3'- and 5' repeat regions of the molecule as well as to non-repeated sequences outside these regions were prepared. T cells from P. falciparum primed donors from two highly endemic areas of Africa were tested for their responsiveness to the peptides by thymidine incorporation and/or interferon gamma (IFN-gamma) release. There was a considerable variation in the response to the different peptides. However, the strongest and most frequent responses were seen with a few peptides from the 3'- and 5'-repeat regions. Thus, the immunodominant B cell epitope regions of Pf155/RESA, contain several T cell epitopes. Since the repeat regions are known to be conserved in different P. falciparum strains, the T cell epitopes reported here may be suitable constituents of a P. falciparum subunit vaccine.

Animals↗
PubMed1988

Monoclonal antibodies to a synthetic peptide corresponding to a repeated sequence in the Plasmodium falciparum antigen Pf155.

Mouse monoclonal antibodies were prepared against a synthetic peptide (EENVEHDA) corresponding to a tandemly repeated sequence in the C-terminus of the Plasmodium falciparum antigen Pf155. One antibody (IgG1) producing hybridoma was studied in detail. The specificity of the antibody was determined by enzyme-linked immunosorbent assays using bovine serum albumin-conjugated or free peptides as solid phase antigens and various synthetic peptides for inhibition. The antibody reacted with Pf155 as detected by immunofluorescence and immunoblotting. It was also an efficient inhibitor of merozoite invasion in P. falciparum in vitro cultures indicating that it defines a biologically important epitope present on the native Pf155 molecule.

Animals↗
PubMed1988

Malaria vaccines: immunogen selection and epitope mapping.

In recent years major efforts have been made to characterize parasite antigens thought to be suitable candidates for malaria vaccines. Many of the relevant plasmodial antigens have been found to contain extensive areas of short amino acid sequences organized in tandem repeats. These are usually strongly antigenic, forming linear epitopes seen by antibodies of the infected host. Several such epitopes have been identified and subunit vaccines are being designed in which synthetic peptides or gene constructs serve as immunogens. However, as an efficient malaria vaccine should give rise to anamnestic T-dependent antibody responses following reinfection after vaccination as well as to antibody independent cell-mediated immunity, efforts are now also being made to identify T-cell epitopes on the vaccine candidate antigens. In this paper the current Plasmodium falciparum sporozoite vaccines and the merozoite antigen Pf155/RESA, a possible candidate for a P. falciparum blood stage vaccine, serve as examples to illustrate recent advances made in this area as well as some of the problems remaining to be resolved.

Animals↗
PubMed1988

T-cell epitopes in Pf155/RESA, a major candidate for a Plasmodium falciparum malaria vaccine.

Immunogens included in a subunit vaccine should contain both B- and T-cell-activating sites to ensure anamnestic responses following reinfection after vaccination as well as antibody-independent cellular immunity. The Plasmodium falciparum antigen Pf155/RESA, a major candidate for a vaccine against the asexual blood stages of this malaria parasite, was investigated for T-cell epitopes in its C-terminal amino acid repeat region, a region known to be conserved in different P. falciparum strains. It was found to contain several related sequences that activated T cells from humans primed to P. falciparum by natural exposure, to proliferation, and/or interferon-gamma release in vitro. T cells from approximately half of the donor group investigated responded to the intact protein, and 65% of these responders also responded to short synthetic peptides, probably representing a small number of partly overlapping T-cell epitopes. Thus, sequences from the C terminus of Pf155 may be suitable constituents of a P. falciparum subunit vaccine and also provide a basis for epitope-specific epidemiological studies relating cellular immune responses in vitro to clinical immunity and P. falciparum endemicity.

Amino Acid Sequence↗
PubMed1986

Human monoclonal antibodies to Pf 155, a major antigen of malaria parasite Plasmodium falciparum.

Pf 155, a protein of the human malaria parasite Plasmodium falciparum, is strongly immunogenic in humans and is believed to be a prime candidate for the preparation of a vaccine. Human monoclonal antibodies to Pf 155 were obtained by cloning B cells that had been prepared from an immune donor and transformed with Epstein-Barr virus. When examined by indirect immunofluorescence, these antibodies stained the surface of infected erythrocytes, free merozoites, segmented schizonts, and gametocytes. They bound to a major polypeptide with a relative molecular weight of 155K and to two minor ones (135K and 120K), all having high affinity for human glycophorin. The antibodies strongly inhibited merozoite reinvasion in vitro, suggesting that they might be appropriate reagents for therapeutic administration in vivo.

Animals↗
PubMed1986

Identification of a Plasmodium chabaudi antigen present in the membrane of ring stage infected erythrocytes.

A novel antigen of asexual blood stages of the rodent malaria parasite Plasmodium chabaudi, was detected by means of a modified indirect immunofluorescence assay (IFA), using glutaraldehyde fixed and air dried monolayers of P. chabaudi infected erythrocytes. P. chabaudi hyperimmune sera gave a distinct surface immunofluorescence of erythrocytes infected with early stages of the parasite. Fixation and drying of the erythrocytes was necessary for the antigenic activity to be exposed. The antigens were species specific as P. chabaudi hyperimmune serum only stained P. chabaudi but not P. yoelii or P. falciparum infected erythrocytes. The antigenic activity involved in the IFA was resistant to trypsin, phospholipases and neuraminidase but not to pronase, suggesting that the antigens were polypeptides. The surface immunofluorescence was inhibited by an extract of parasitized erythrocytes, but not by similar extracts of normal erythrocytes. The inhibitory antigens were soluble and heat stable (100 degrees C, 5 min). For identification and characterization of the antigens, antibodies were isolated by acid elution from monolayers of infected erythrocytes and monoclonal antibodies were produced. Probing in immunoblotting of extracts of parasitized erythrocytes separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis with the eluted antibodies, showed that they reacted consistently with a polypeptide of Mr 105 000 (Pch105). The Pch105 antigen shares many characteristics with Pf155, a P. falciparum antigen considered as a candidate for a vaccine against that parasite.

Animals↗
PubMed1986

Rabbit and human antibodies to a repeated amino acid sequence of a Plasmodium falciparum antigen, Pf 155, react with the native protein and inhibit merozoite invasion.

The Plasmodium falciparum-derived antigen of Mr 155,000 designated Pf 155, deposited in the membrane of infected erythrocytes, contains at least two blocks of tandemly repeated amino acid sequences. The peptide Glu-Glu-Asn-Val-Glu-His-Asp-Ala, which corresponds to a subunit of a C-terminally located repeat, was synthesized. Rabbits immunized with the octapeptide conjugated with either keyhole limpet hemocyanine or tetanus toxoid formed antibodies against the octapeptide. These antibodies reacted with Pf 155 as detected by immunoblotting or a modified immunofluorescence assay. Sera from humans exposed to P. falciparum also contained antibodies binding to the octapeptide in a dot-blot immunoassay. Their anti-octapeptide titers were correlated with their immunofluorescence titers in the assay detecting Pf 155 and other parasite antigens in the membrane of infected erythrocytes. Human octapeptide-reactive antibodies were isolated on an affinity column with the octapeptide conjugated to bovine serum albumin as ligand. These human antibodies reacted with Pf 155 in immunoblotting and strongly stained the surface of infected erythrocytes in the modified immunofluorescence assay. Approximately 20% of this immunofluorescence activity in a high-titered human serum could be recovered from the octapeptide column, indicating that a significant fraction of these anti-parasite antibodies react with epitopes associated with the octapeptide. Furthermore, the human octapeptide-reactive antibodies very efficiently inhibited merozoite reinvasion into erythrocytes in vitro. Similarly purified rabbit antibodies also significantly inhibited reinvasion. Our results suggest that the C-terminal segment of repeated peptides in Pf 155 is a major antigenic region of the molecule and may contain target sites for protective immunity in P. falciparum malaria.

Amino Acid Sequence↗
PubMed1986

Anti-Plasmodium falciparum antibodies acquired by residents in a holoendemic area of Liberia during development of clinical immunity.

Sera from 48 children and adolescents (2-15 years of age), residing in a malaria holoendemic area of Liberia were investigated for specificities and isotypes of anti-P. falciparum antibodies. No clear-cut relationship to the development of clinical immunity was found when the overall antibody activities to total parasite antigens were determined by enzyme-linked immunosorbent assay (ELISA). Although there was a certain rise of IgM, total IgG- and IgG2 antibody activities, this was most pronounced at ages when a clinical but nonsterile immunity is already present. When the sera were investigated by immunoprecipitation of 35S-methionine labeled parasite polypeptides, the total number of parasite antigens precipitated was similar at all ages. Analysis by indirect immunofluorescence (IFA), registering antibodies to intracellular parasite antigens, revealed no age-dependent changes in antibody titers. In contrast, when the sera were assayed by a novel IFA, specific for a restricted number of parasite antigens in the membrane of infected erythrocytes, the frequency of positive sera as well as the anti-P. falciparum titers rose in parallel with the development of clinical immunity. Thus, these antigens appeared to be important inducers of protective immune responses and may be suitable candidates for a vaccine against the asexual blood stages of P. falciparum.

Adolescent↗
PubMed1986

Characterization of the humoral immune response in Plasmodium falciparum malaria. III. Factors influencing the coexpression of antibody isotypes (IgM and IgG-1 to 4).

Isotypes (IgM and IgG-1 to 4) of anti-P. falciparum antibodies were investigated in sera of malarial patients or immune donors by enzyme linked immunosorbent assay (ELISA) and two indirect immunofluorescence assays (IFAs), one staining intra-erythrocytic parasites of all stages and the other a restricted number of parasite antigens deposited in the membrane of infected erythrocytes by invading merozoites (Perlmann & Wahlgren, 1983; Perlmann et al., 1984, Wahlgren et al., 1985a). There was no correlation in overall antibody titres between the two IFAs. Antibodies of both IgM and all four IgG isotypes were detected in both assays. With the IFA for intracellular parasites a brilliant fluorescence was obtained with antibodies of all isotypes. However IgG-2 antibodies often gave staining restricted to the surface of schizonts. The incidence and reactivity in individual sera of antibodies of the different isotypes did not relate to the immune status of the donors (acute infection or clinically immune) but related well to the degree of malarial exposure as reflected by the overall antibody titres. This, in all three assays, high titred sera frequently contained antibodies of all isotypes while low titred sera usually only contained antibodies of IgM, IgG-1 and IgG-3 isotype. On average, the overall expression of antibodies of different isotypes in individual sera appears to reflect a sequential downstream (5' to 3') activation of the corresponding Igh-C genes in P. falciparum specific B-cell clones.

Adolescent↗
PubMed1985

Pf 155, a candidate for a blood stage vaccine in Plasmodium falciparum malaria.

Pf 155 is a Mr 155,000 P. falciparum antigen, which is deposited in the erythrocyte membrane at merozoite invasion. The antigen is detected by a modified immunofluorescence assay giving staining of the surface of ring stage infected erythrocytes. Cell lines producing human monoclonal antibodies to Pf 155 were established and two different antibodies of IgM and IgG class, respectively, were characterized further. Both antibodies gave a strong surface immunofluorescence and in immunoblotting they react strongly with Pf 155, but also with some lower molecular weight material, including polypeptides of Mr 135,000 and 120,000. Both antibodies were efficient inhibitors of P. falciparum reinvasion in vitro. Pf 155 as well as the Mr 135,000 and 120,000 polypeptides were shown to bind with high affinity to human erythrocyte glycophorin. An octapeptide (GluGluAsnValGluHisAspAla) corresponding to a repeated sequence in Pf 155 was synthesized. Rabbit antibodies to the octapeptide gave a distinct surface immunofluorescence and reacted with Pf 155 and the Mr 135,000 and 120,000 polypeptides in immunoblotting. Human antibodies reacting with the octapeptide were isolated by affinity chromatography from the serum of a P. falciparum immune individual. These antibodies showed a strong reaction with Pf 155 as determined by immunoblotting and surface immunofluorescence. Pf 155 reactive antibodies affinity purified on monolayers of P. falciparum infected erythrocytes are very efficient inhibitors of parasite reinvasion. Such antibody preparations depleted of octapeptide reactive antibodies showed a markedly decreased reinvasion inhibitory capacity, while a high inhibitory activity was recovered in the octapeptide reactive antibodies. Pf 155 fulfills several criteria thought to be proper for antigens involved in anti-malarial protective immunity.

Amino Acid Sequence↗
PubMed1984

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

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)

Antibodies↗