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Biomedical subjects

G F Mitchell

Publications and source records attributed to G F Mitchell.

At least 181 records · Page 10Linked to original sources
PubMed1981

Hybridoma-derived antibody with immunodiagnostic potential for schistosomiasis japonica.

A murine hybridoma-derived antibody (IPH.134) has been produced which has apparent high binding specificity for an extract of Schistosoma japonicum adult worms. No binding was detected to extracts of S. japonicum eggs or to extracts from other adult trematodes (Fasciola hepatica, Paragonimus westermanii, Clonorchis sinensis, and Schistosoma mansoni) and several other helminths and protozoa. When sera from two series of S. japonicum-infected Philippine patients (19 and 20 patients, respectively) were tested for inhibition of binding of 125I-labeled IPH.134 to S. japonicum adult worm antigen, a low (10%) false-negative rate was obtained. In the 19-patient series, the 4 patients with highest fecal egg counts had high inhibitory activity in their sera. In the 20-patient series, the 3 patients with prominent disease had high inhibitory activity in their sera. Evidence was obtained that IgG anti-S. japonicum antibodies (rather than circulating antigens, immune complexes, or anti-idiotypic antibodies) were most likely to be responsible for serum inhibitory activity in the test. No false-positive reactions have been obtained with pooled or individual sera from patients infected with numerous parasites other than S. japonicum, although no information is yet available on the inhibitory activities of sera fro S. mansoni- or S. hematobium-infected individuals. On the basis of the data obtained to date, it is a reasonable prediction that the molecule or determinant to which this hybridoma antibody is directed will be a useful immunodiagnostic antigen for schistosomiasis japonica in the Philippines. A test based on detection of serum antibodies to this antigen should have high specificity and may provide additional information on the level of infection or disease status in patients.

Adolescent↗
PubMed1981

Immunoprecipitation of biosynthetically-labelled proteins from different Papua New Guinea Plasmodium falciparum isolates by sera from individuals in the endemic area.

The human serum antibody response to Plasmodium falciparum infection in Papua New Guinea has been studied by electrophoretic analysis of immunoprecipitated biosynthetically-labelled malaria proteins from three different isolates maintained in long-term in vitro culture. Differences in protein antigenic composition in different lines have been described and simplified by examination of antigens recognized only by hyperimmune serum. An in vitro assay has been used to screen various human sera containing antimalarial antibody for their ability to inhibit parasite growth and the immunoprecipitation profiles of non-inhibitory sera have been compared with those of a hyperimmune serum pool. In the discussion, emphasis is placed on the value of immunoprecipitation analyses using clinically-defined sera with known in vitro function in the identification of antigens which may be responsible for the induction of host-protective immunity.

Adult↗
PubMed1981

Clonal parasites in the analysis of resistance to reinfection with Fasciola hepatica.

A feature of fascioliasis in mice and rats is incomplete and generally poor resistance to reinfection. The possibility exists that population heterogeneity amongst Fasciola hepatica parasites (in characteristics such as antigenic variability and infectivity) contributes to the incomplete resistance expressed by already infected rodents to challenge infection. Using different exposure regimes, mice and rats were dosed with infective metacercariae of different single snail-derived clones and challenged with the same or different clonal parasites. The results clearly demonstrate that no better resistance to reinfection is seen with parasites of the homologous clone than with heterologous clone challenge. Thus the poor resistance to reinfection seen in fascioliasis cannot be ascribed readily to antigenic or infectivity differences between clonal metacercariae.

Animals↗
PubMed1981

Identification of radioiodinated cuticular proteins and antigens of Onchocerca gibsoni microfilariae.

The filarial parasite of cattle, Onchocerca gibsoni, has been used to establish procedures of antigen identification with a view of applying these techniques to studies on human filarial parasites. Emphasis has been placed on methods suitable for use with small numbers of parasites. Microfilariae (mf) of O. gibsoni were extracted from nodular worms, purified and 125I-labeled using IODO-GEN in solid-phase. Radioactivity was shown to be confined to the cuticle of sectioned mf using the technique of electronmicroscope autoradiography. Radiolabeled mf were analysed by two-dimensional gel electrophoresis. Autoradiographs of 125I-labeled proteins of O. gibsoni mf were relatively complex, there being at least 32 proteins ranging in molecular weights from 20,000 to 120,000 and displaying considerable charge heterogeneity. Evidence was obtained that at least the major serum proteins of the host, albumin or immunoglobulin, were not absorbed on the surface of these uterine mf and detectable in the labeled surface protein patterns. Sera from infected cattle immunoprecipitated 5 labeled proteins from a Triton X-100 extract of 125I-labeled mf. Sera from either of two calves which had been given multiple injections of mf subcutaneously, and which had no detectable skin mf, recognised 6 additional proteins in this extract as well as 3 of the proteins recognised by sera from infected cattle.

Animals↗
PubMed1981

Hybridoma antibody immunoassays for the detection of parasitic infection: further studies on a monoclonal antibody with immunodiagnostic potential for Schistosomiasis japonica.

The binding of a hybridoma-derived antibody (designated IPH.134-18-6) to an extract of the adult worn of Schistosoma japonicum has been further characterized. This antibody has immunodiagnostic potential for detection of infection with S. japonicum in the Philippines since the binding of labeled hybridoma antibody to a crude adult worm extract in a solid-phase radioimmunoassay (RIA) is inhibited by sera from greater than 90% of known infected individuals, and no false positive reactions have yet been observed. Moreover, most patients with high fecal egg outputs or prominent disease have high serum inhibitory activity. All evidence indicates that IgG antibodies are the serum inhibitors in the competitive RIA. By estimating the amount of inhibitory activity (antibody?) of IPH.134-18-6 type in sera of infected patients and animals, the conclusion was reached that the antigen to which this hybridoma is directed is a strong immunogen. Moreover, no evidence of genetic unresponsiveness was obtained in strains of inbred mice infected with S. japonicum. IPH.134-18-6 does not bind to S. japonicum egg extracts. However, the actual stage and species specificity of this antibody has not yet been determined unequivocally and must await the screening of sera from individuals with monospecific S. mansoni and S. hematobium infections as well as assays using schistosomules, male and female adult worms and juveniles.

Animals↗
PubMed1981

The major radioiodinated cuticular antigens of Onchocerca gibsoni microfilariae are neither species nor onchocerca specific.

The possible role of microfilarial surface (cuticular) antigens in immuno-diagnosis of human filarial infections has been assessed using microfilariae (mf) of the cattle parasite Onchocerca gibsoni. A Triton X-100 extract of 125I-labeled O. gibsoni mf was reacted with a panel of sera from humans infected with Onchocerca volvulus, Wuchereria bancrofti and Schistosoma japonicum as well as sera from uninfected controls. Results of these immunoprecipitations indicated that sera from humans infected with O. volvulus or W bancrofti contain antibody specificities recognising certain of the radioiodinated cuticular proteins of O. gibsoni mf. Two-dimensional gel analysis and subsequent autoradiography of these immunoprecipitates showed that 8 radioiodinated proteins recognised by sera from calves injected with O. gibsoni mf were also immunoprecipitated by sera from humans infected with either O. volvulus or W. bancrofti. Thus there appear to be no major radioiodinated cuticular antigens of O. gibsoni mf which are species or onchocerca specific.

Animals↗
PubMed1981

Schistosoma japonicum: use of a radioimmunoassay for anti egg antibodies in human sera.

A solid-phase radioimmunoassay (RIA) was developed for schistosomiasis japonica using extracted egg antigens and compared with circumoval precipitin test (COPT) results on 20 sera from known S. japonicum-infected individuals and on 10 control sera. The quantitative RIA very clearly differentiated between infected and uninfected individuals with highest titers being obtained in teenagers. However, in the series employed, information relevant to immunodiagnosis of S. japonicum infection was contained in the non-quantitative but simple COPT and little was apparently added to the quantitative but expensive RIA.

Antibodies↗
PubMed1981

Attempts to standardize the circumoval precipitin test (copt) for Schistosomiasis japonica.

The circumoval precipition test (COPT) is a simple and inexpensive immunodiagnostic test for schistosomiasis japonica which, in the Philippines, has high sensitivity and specificity. Lack of standardization does, however, increase the variability of the test. Parameters which influence the COPT have been examined using large numbers of sera from known S. japonicum infected individuals. In this series of experiments, optimal conditions were determined to be as follows using 2 drops of neat serum and incubation at 37 degrees C in a sealed slide chamber; - approximately 100 eggs from 55 or 60 days infected rabbits for a 24 to 48 hour incubation period. COP reactions (i.e. precipitates associated with eggs) were much less obvious when either immature eggs or eggs obtained from long-term infected rabbits were used. The results emphasize the prime importance of the source of Schistosoma japonicum eggs in the performance of a standardized COPT.

Animals↗
PubMed1981

Identification and characterization of protein antigens of Leishmania tropica isolates.

Promastigotes of 4 Leishmania tropica isolates were biosynthetically labeled with 35S-methionine or surface radioiodinated, and the detergent lysates were analyzed by 2 dimensional gel electrophoresis. The protein patterns of cytoplasmic and membrane proteins detected in two independent isolates from simple cutaneous leishmaniasis cases were similar to each other, but were different from 2 isolates taken over a 20-yr interval from a case of leishmaniasis recidiva. The analysis of radioiodinated membrane proteins of L. tropica promastigotes revealed a simple pattern of 6 to 7 labeled major proteins and some minor ones. Unlike cytoplasmic proteins, membrane proteins were highly resistant to digestion by proteases. Partial cleavage of radioiodinated surface proteins by staphylococcal V-8 protease in the presence of 4.5 M urea and peptide mapping confirmed the presence of shared and isolate-specific proteins. The significance of these findings for the classification of Leishmania spp. and the pathogenesis of the different disease states that they cause are discussed.

Animals↗
PubMed1981

Giardiasis in mice: studies on the characteristics of chronic infection in C3h/He mice.

When C3H/He mice are dosed orally with fecal cysts or trophozoites of the intestinal parasite, Giardia muris, they become chronically infected, whereas BALB/c (or DBA/2) mice rapidly eliminate their infection. The ability to eliminate a primary infection or resist a challenge infection was not related to levels of serum IgA antibody directed against a crude extract of trophozoites. The possibility that the development of host-protective immunity may either be delayed or absent in C3H/He mice was investigated by 2 types of experiment: 1) BALB/c and C3H/He mice were drug-cured at various times after a primary infection and assessed for their ability to resist reinfection, and 2) the ability of infected female mice to protect their suckling young was assessed. Under these experimental conditions, no significant difference in the kinetics of development of resistance to a challenge infection was noted. Thus, mice of both strains were capable of resisting a challenge infection after drug-cured and infected mothers were able to protect their suckling young from a challenge infection. Drug-cured long-term infected mice were resistant to a challenge of either cysts or trophozoites obtained from either 1-wk infected nude mice or long-term infected C2H/He mice. Thus, it is unlikely that the explanation for chronic infection oif C3H/He mice is that variants or a modified population of G. muris trophozoites arise to which C3H/He mice have a defective response. The possibility that G. muris is able to suppress host-protective responses in chronically-infected C3H/He mice but not in BALB/c mice is discussed.

Animals↗
PubMed1980

Characterization of surface proteins and glycoproteins on red blood cells from mice infected with haemosporidia: Babesia rodhaini infections of BALB/c mice.

Infection of intact (nu/+) or hypothymic (nu/nu) BALB/c mice with the lethal intra-erythrocytic parasite, Babesia rodhaini, induced several changes in the surface proteins of red cells from infected blood. Lactoperoxidase-catalysed radio-iodination was used to compare the surface proteins on normal mouse erythrocytes and the total red cells from infected blood at different levels of parasitaemia. At very low parasitaemia, when only 2.5% of the red cells contained parasites, we observed significant changes in the profile of radio-iodinated proteins separated by SDS-polyacrylamide gel electrophoresis. These changes included the appearance of a group of high molecular weight proteins, and a protein with an apparent molecular weight (Mr) of 60 000, both of which were absent from normal erythrocytes. The major labelled band on the erythrocyte surface (Mr 92 000) also appeared to be less heavily labelled during infection. The magnitude of these differences in surface proteins increased as the parasitaemia rose, until the new bands dominated the radioactivity profile with blood of greater than 50% parasitaemia. Several control experiments established that the radio-iodinated proteins were surface molecules on intact cells and that artifactual proteolysis did not contribute to the observed differences. The results suggest that changes in the surface proteins occur on all red cells in the blood of infected mice. The results of labelling the surface glycoproteins by oxidation with periodate or galactose oxidase, followed by NaB3H4 reduction, have varied with the isolate of B. rodhaini. With the isolate currently in use, no significant differences were observed in the labelled surface glycoproteins of normal erythrocytes and red cells from infected blood of high parasitaemia, whereas an earlier isolate exhibited a marked decrease of glycoprotein labelling of both infected and uninfected red cells.

Animals↗
PubMed1980

Characterization of surface proteins and glycoproteins on red blood cells from mice infected with haemosporidia: Plasmodium berghei infections of BALB/c mice.

The surface proteins and glycoproteins of red cells from Plasmodium berghei-infected blood have been radio-isotope labelled and compared with those of normal mouse erythrocytes using the following protein labelling probes: lactoperoxidase-catalysed radio-iodination of tyrosyl residues, periodate oxidation and NaB3H4 reduction of sialic acid and oxidation of galactosyl/N-acetylgalactosaminyl residues by galactose oxidase with subsequent NaB3H4 reduction. During P. berghei infection, new tyrosyl-labelled proteins with apparent molecular weights (Mr) of 60 000, 54 000, 40 000 and 27 500 appeared on the surface of most, if not all, red cells in the blood. Purified multinucleate cells (mostly reticulocytes) differed only in that they also had a surface protein with Mr of 83 000. However, this molecule is thought to be specific to mouse reticulocytes rather than derived from parasites. In contrast to the relatively minor changes detected with radio-iodination, striking changes in glycoprotein radio-isotope labelling resulted from infection. All of the red cells in infected blood of greater than 20% parasitaemia lost their periodate-sensitive glycoprotein sialic acid. With some samples there was little change in glycoprotein labelling by the galactose oxidase method, provided neuraminidase was also added. Modification of the exocyclic hydroxyls of sialic acid is postulated to account for this. Other blood samples exhibited a dramatic loss of galactose oxidase-dependent labelling. It is suggested that these observations may relate to the excessive red cell destruction of uninfected as well as infminidase was also added. Modification of the exocyclic hydroxyls of sialic acid is postulated to account for this. Other blood samples exhibited a dramatic loss of galactose oxidase-dependent labelling. It is suggested that these observations may relate to the excessive red cell destruction of uninfected as well as infminidase was also added. Modification of the exocyclic hydroxyls of sialic acid is postulated to account for this. Other blood samples exhibited a dramatic loss of galactose oxidase-dependent labelling. It is suggested that these observations may relate to the excessive red cell destruction of uninfected as well as infected cells which has been inferred in many haemosporidial infections, including malaria.

Animals↗
PubMed1980

Characterization of surface proteins and glycoproteins on red blood cells from mice infected with haemosporidia: Plasmodium yoelii infections of BALB/c mice.

Lactoperoxidase-catalysed radio-iodination was used to compare the surface proteins on red cells from Plasmodium yoelii-infected with normal BALB/c mice. The profile of radio-iodinated proteins separated by SDS-polyacrylamide gel electrophoresis was different for infected blood of similar parasitaemia from mice inoculated with different doses of the parasite. Inoculation with different doses of the parasite. Inoculation with the lower dose resulted in the appearance of a major radio-iodinated protein of apparent molecular weight (Mr) 76 000 which was labelled to a similar extent on uninfected red cells from infected blood and purified multinucleate infected cells. Several minor radio-iodinated bands, with identical mobilities to the minor bands on normal BALB/c erythrocytes, were also present on red cells from this infected blood. In contrast, the higher inoculation dose produced changes in the minor labelled bands, and the band with Mr of 76 000 was absent. In this case, the minor radio-iodinated proteins of the normal BALB/c erythrocyte (with Mr of 65 000, 57 000, 48 000, 38 000 and 32 000) were replaced by a series of bands with Mr of 60 000, 50 000, 43 000 and 28 000 on both uninfected and infected red cells. These differences with inoculation dose may be related to the different duration of these infections, the development of anaemia and the extent of pathological changes at the erythrocyte surface. P. yoelii infection caused a marked loss in periodate-dependent labelling of sialoglycoproteins on most, if not all, red cells in infected blood. There was also a large decrease in galactose oxidase-dependent glycoprotein labelling with or without neuraminidase treatment. These changes in the carbohydrate groups on red cell membrane glycoproteins may be linked to the excessive loss of both uninfected and infected red cells during some malaria infections.

Animals↗
PubMed1980

A difference in surface proteins of Fasciola hepatica larvae from intact and nude mice.

Larvae of the trematode, Fasciola hepatica, dissected from the livers of intact mice at 12 to 14 days of infection, bind anti-immunoglobulins in vitro. This is not so for larvae harvested from the livers of infected hypothymic nude (nu/nu) mice. When attempts were made to confirm the presence of surface Ig using lactoperoxidase-catalysed radioiodination and analysis by SDS-polyacrylamide gel electrophoresis, proteolysis at the time of solubilization markedly affected the profiles obtained. Provided protease inhibitors were present during processing, a mu-like Ig heavy chain and a gamma-like Ig heavy chain could be demonstrated on larvae from nu/+ mice but not on larvae from nu/nu mice.

Animals↗