Chev Kidson

A rhoptry antigen of Plasmodium falciparum contains conserved and variable epitopes recognized by inhibitory monoclonal antibodies.

Four monoclonal antibodies produced against Plasmodium falciparum recognize an antigen in merozoites that is localized in rhoptries, as judged by a punctate, double dot fluorescence pattern. All four antibodies bound to the same affinity purified antigen in a two site immunoradiometric assay. Immunoprecipitation of antigen by monoclonal antibody followed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis yielded protein bands of 80, 66 and 42 kDa. Western blotting gave bands of 80 and 66 kDa only with three of the antibodies: the fourth did not blot. Based on protease inhibitor data the 66 kDa band is considered to be a cleavage product of the 80 kDa band, but the 42 kDa band does not appear to derive from the latter and may be a coprecipitation product. This group of antigens labels with both [35S]methionine and [3H]histidine. Two of the monoclonal antibodies inhibited merozoite invasion of erythrocytes. One of these inhibitors recognizes a variable epitope, whereas the second recognizes a highly conserved epitope present in all 106 primary isolates of P. falciparum tested from Brazil, Thailand and Papua New Guinea.

Anti‐mosquito antibodies decrease the reproductive capacity of Aedes aegypti

Abstract. Aedes aegypti (L.) fed on rabbits immunized with mosquito antigens showed a reduction in fecundity in the first oviposition and decreased viability of the progeny. Feeding behaviour of mosquitoes was not affected and no significant mortality was observed due to the presence of anti‐mosquito antibodies in the bloodmeal. Antibodies were detected in the oocytes of mosquitoes 48 h after the bloodmeal. The role of specific antibodies in influencing fecundity is discussed.

A high molecular weight antigen in Plasmodium falciparum recognized by inhibitory monoclonal antibodies

Summary Inhibitory monoclonal antibodies which bind to some isolates of Plasmodium falciparum from Papua New Guinea, but not from other areas, bound to a 220 kD antigen. By immunofluorescence microscopy this antigen was shown to be located both within the schizont cytoplasm and also within the schizont infected erythrocyte, but external to the schizont itself. Even at antibody concentrations which caused > 70% inhibition of parasite multiplication, accumulation of schizont stages or aggregates of merozoites were not seen, consistent with inhibition occurring at a point after the release of merozoites. While this suggests that the antigen may be present on merozoites, the quantity was below the limit of detection. It is suggested that the large amount of antigen released by rupturing schizonts may be a mechanism used by the parasite to evade immunological attack.

Chemical characterization of the parasitophorous vacuole membrane antigen QF 116 from Plasmodium falciparum

On the basis of amino acid sequencing and immunological cross-reactivity, the Plasmodium falciparum parasitophorous vacuole antigens QF116 and exp-1/CRA are apparently identical. The epitope recognized by an inhibitory monoclonal antibody directed against QF116 is located proximal to the C-terminus of the protein. The QF116 protein is processed during maturation by the cleavage of a 22-amino-acid signal peptide and acylated as measured by labeling with myristic acid.

Perturbations of cell-cycle progression in γ-irradiated ataxia telangiectasia and Huntington's disease cells detected by DNA flow cytometric analysis

The effects of ionizing radiation on cell-cycle progression in lymphoblastoid cell lines derived from ataxia telangiectasia (AT) and Huntingtons disease (HD) patients, and from normal individuals, were studied using DNA flow cytometric analysis. A dose of 100 rad gamma irradiation blocked a proportion of normal and HD cells in G1. A higher radiation dose applied to normal cells increased the number of cells blocked in G1 and significantly delayed cells which were in S at the time of irradiation from reaching G2 DNA content. The reduced cumulative mitotic index in irradiated cultures of normal cells 2 h after irradiation suggests that cells in G2 at the time of irradiation are delayed before entering mitosis. After irradiation HD cells responded similarly to normal cells except that a greater proportion of HD cells were blocked in G1. AT cells do not show the normal delay in progression from G1 to S, or from S to G2 in the first cycle after irradiation. The cumulative mitotic index was reduced in irradiated cells, implying that they are delayed in G2. Thus AT cells did not recognize or respond to signals from damaged DNA which in normal and HD cells caused a proportional block in G1 and an S-phase delay. The only point of arrest in cell-cycle progression in irradiated AT cells was in G2.

Gene dosage and complementation analysis of ataxia telangiectasia lymphoblastoid cell lines assayed by induced chromosome aberrations

An approach of general applicability to mammalian radiosensitive mutants has been used in the analysis of gene dosage and complementation in ataxia telangiectasia (A-T). Thymidine residues in DNA of one parental lymphoblastoid cell line were substituted with bromodeoxyuridine before fusion with a second parental cell line, to allow differential staining of the two sets of chromosomes. Following gamma-irradiation, induced chromosome aberrations were scored in diploid and homokaryon cells from each parental line as well as in heterokaryons. Four complementation groups were ascertained among 7 A-T cell lines. Analysis of heterokaryons formed between appropriate combinations of normal, A-T homozygote and A-T heterozygote cells, gave a quantitative measure of gene dosage and demonstrated increasing radiosensitivity with increasing numbers of A-T alleles.

Huntington's disease: implications of associated cellular radiosensitivity

Ionizing radiation sensitivity was studied in a series of Huntingtons Disease (HD) patients and controls by measurement of radiation‐induced chromosome aberrations in lymphocytes and by clonogenic survival of lymphoblastoid cell lines. As a group, HD patients were found to be significantly more radiosensitive than controls (p < 0.001), but there was an overlap between values for the two groups such that an absolute distinction is not possible. These data are consistent with an association between HD and radiosensitivity but not with identity between HD and a radiosensitive phenotype, so that cellular radiosensitivity cannot be used for individual diagnosis. Analysis of three families including 5 HD patients and 11 first‐degree relatives confirmed this conclusion and demonstrated that even within a given family presymptomatic diagnosis cannot be based on measurement of radiosensitivity. However, the common association of cellular radiosensitivity with HD probands and their families provides a potential lead to the identification of HD gene(s) and so to an eventual understanding of the aetiopathogenesis of this disease at the molecular level.

Plasmodium falciparum: Automated assay of erythrocyte invasion using flow cytofluorometry

Abstract Erythrocytes labeled with fluorescein isothiocyanate, were mixed with erythrocytes infected with Plasmodium falciparum . After allowing time for invasion of labeled cells to take place, cells were stained with propidium iodide. Parasitemia in labeled cells was determined using flow cytofluorometry. The invasion of labeled erythrocytes was reduced in a dose-dependent manner by immune serum. The degree of inhibition obtained increased as the erythrocyte concentration decreased.

Plasmodium falciparum: Assay of invasion of erythrocytes

Abstract A method for quantitatively assaying Plasmodium falciparum merozoite invasion of particular erythrocytes is described. Erythrocytes were labeled with fluorescein isothiocyanate which did not affect parasite entry or growth, to distinguish them from uninfected erythrocytes in the original parasitized cell population. Parasites were detectable after staining with ethidium bromide. The time course of infection of the labeled cells was followed over 26 hr. The technique was used to determine the effect of serum from a patient with P. falciparum malaria on merozoite invasion of the labeled erythrocytes.

The synthesis and fate of stage-specific proteins in Plasmodium falciparum cultures

Cultured ring, trophozoite and schizont stages of Plasmodium falciparum were metabolically labeled with [35S]methionine. After labeling, cultures were incubated for varying times in the presence of non-radioactive methionine. Triton-soluble proteins from different stages of growth were analysed by sodium dodecylsulfate-polyacrylamide gel electrophoresis. Most proteins were synthesized by every stage of growth and remained unchanged throughout the cycle through to the ring stage following merozoite invasion of erythrocytes. At least 15 proteins, most of high molecular weight, were synthesized solely or predominantly by schizonts. Eight proteins (approx. 177, 170, 158, 87, 83, 47, 41 and 24 kDa) appeared in schizonts but not merozoites. Eight proteins (approx. 240, 203, 106, 80, 35, 19, 15 and 14 kDa) appeared in merozoites, but not in rings following merozoite invasion. Some proteins appeared to be modified after synthesis.