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Biomedical subjects

L E Brown

Publications and source records attributed to L E Brown.

At least 73 records · Page 4Linked to original sources
PubMed1986

Antigenic determinants of influenza virus hemagglutinin. XII. the epitopes of a synthetic peptide representing the C-terminus of HA1.

A synthetic peptide comprising the C-terminal 24 amino acids of the heavy chain (HA1) of influenza virus hemagglutinin was constructed and examined for antigenic and immunogenic activity. Monoclonal antibodies as well as polyclonal antisera raised against the synthetic peptide were able to bind to intact virus. This binding was greatly enhanced if the virus was first subjected to pH 5, suggesting that this treatment exposes the C-terminus of HA1. Using synthetic analogs of the native sequence it was shown that the epitope recognized by one of the monoclonal antibodies encompasses one or more of the C-terminal four amino acids of HA1 (residues 325-328), which are conserved within subtypes but differ between subtypes, while the other monoclonal antibody recognizes a different epitope which involves at least one of the five variable residues at positions 311-315.

Amino Acid Sequence↗
PubMed1985

Location of antigenic sites on the three-dimensional structure of the influenza N2 virus neuraminidase.

Sequence analysis of the neuraminidase (NA) genes of influenza virus X-7(F1) and of 12 variants selected with monoclonal antibodies has been used to define in physical terms the antigenic structure of this NA, which was operationally established by R. G. Webster, L. E. Brown, and W. G. Laver (1984, Virology 135, 30-42). X-7(F1) is a reassortant virus containing the NA of the early Asian (H2N2) isolate A/RI/5+/57, and the results of antigenic and sequence analysis of X-7(F1) and of variants selected with monoclonal antibodies have been combined with a similar analysis of the A/Tokyo/3/67 NA (H2N2, M. R. Lentz, G. M. Air, W. G. Laver, and R. G. Webster (1984), Virology 135, 257-265) to obtain a model of antibody binding to N2 NAs. The selection process was biased, however, since only those monoclonal antibodies which inhibited NA activity could be used to select variants. Most of the changes in the variants selected with monoclonal antibodies occur in those parts of the polypeptide chain which encircle the enzyme active site pocket in the three-dimensional structure (P. M. Colman, J. N. Varghese, and W. G. Laver (1983), Nature (London) 303, 41-44). The results suggest that in general the antibody binds to a site on the NA which includes those amino acid side chains which are altered in monoclonal variants. There are, however, several aspects of the antigen-antibody interaction which are not easily explained, and which will probably only be fully elucidated by X-ray crystallographic analysis of NA-antibody complexes.

Amino Acid Sequence↗
PubMed1985

Subcellular distribution of [125I]iodoaryl beta-methyl fatty acids.

Subcellular distribution studies of two aryl branched-chain and one aryl straight-chain iodinated fatty acids were carried out as part of a continuing effort to determine if such acids are metabolized by beta-oxidation. For the omega-iodoaryl fatty acids, a change in subcellular radioactivity location was observed which was chain-length dependent. Chain lengths of 15 carbons, straight and branched, were largely found in the nuclear-membrane fraction, whereas a chain length of 8 carbons was largely located in the cytosol. No unequivocal evidence for metabolic trapping in the mitochondria was observed for omega-iodoaryl branched-chain fatty acids using the centrifugation technique employed in this study.

Animals↗
PubMed1985

Antigenic analysis of H2 influenza virus haemagglutinin with monoclonal antibodies.

Antigenic analysis of human and avian H2 influenza virus was carried out with monoclonal antibodies to the HA molecules of H2 influenza viruses isolated in the early stage of an H2 pandemic. The study revealed antigenic differences between inhibitor sensitive (Japan+/57, RI+57) and inhibitor resistant strains (Japan-/57, Ri-/57). This indicates that the receptor-binding specificity of the haemagglutinin can markedly influence the antigenic analysis obtained with monoclonal antibodies in HI test. Minor antigenic differences (microheterogeneity) could be detected between different H2 influenza viruses isolated in 1957. Minor antigenic variation continued in the H2 viruses until 1961, but significant antigenic drift occurred in 1962 so that viruses isolated after that date reacted with few monoclonal antibodies. Analysis of avian H2 influenza viruses suggested antigenic differences between the different avian H2 haemagglutinin, but no correlation between the year of isolation and the progressive antigenic drift similar to that seen in the human strains was found.

Antibodies, Monoclonal↗
PubMed1985

Murine helper T lymphocyte response to influenza virus: recognition of haemagglutinin by subtype-specific and cross-reactive T cell clones.

Helper T cel lines specific for influenza virus were established by continuous culture of Mem 71-Bel (H3) virus-immune spleen cells in the presence of virus and antigen-presenting cells and their specificity assessed in proliferation experiments. A line stimulated in vitro with Mem 71-Bel virus was able to proliferate in response to viruses of the same, and also of different, type A haemagglutinin (HA) subtypes as the immunizing virus but not to a type B influenza virus. A component of this cross-reactivity was due to recognition of the HA molecule. Lines stimulated in vitro with purified disrupted H3 or H2 viruses showed a higher degree of cross-reactivity. Of nine clones isolated from these lines, seven were directed against the HA molecule and recognized the HA1 chain. The HA-specific T cell clones were either subtype-specific T cell clones were either subtype-specific, recognizing only viruses of the H3 subtype, or cross-reactive, also recognizing viruses of the H2 subtype of type A (but not type B). Subtype-specific and cross-reactive T cell clones were shown to function as helper T cells in vitro. In addition to collaborating with H3 virus-primed B cells responding to H3 virus in culture, the cross-reactive T cell clone could also provide help for H2 virus-primed B cells making anti-HA antibody in response to H2 virus.

Animals↗
PubMed1984

Characterization of H2 influenza virus hemagglutinin with monoclonal antibodies: influence of receptor specificity.

Antigenic analysis of human and avian H2 influenza viruses were done with monoclonal antibodies to the HA molecules in hemagglutination inhibition (HI) assays. These studies revealed that the receptor-binding specificity of the hemagglutinin can markedly influence the antigenic analysis obtained with monoclonal antibodies in HI tests. Influenza viruses that are sensitive or resistant to inhibition by horse serum inhibitors showed marked differences in their reactivity with monoclonal antibodies to the hemagglutinin. This was apparent with the A/RI/5+/57 and A/RI/5-/57 strains of H2N2 viruses isolated by Choppin and Tamm (1960a), half of the panel of different monoclonal antibodies failed to inhibit hemagglutination of the RI/5- variant, whereas all of the 18 monoclonal antibodies inhibited RI/5+. These findings have important implications in the antigenic analysis of influenza viruses where HI assays are conventionally used to determine the extent of antigenic drift in nature. Antigenic differences were detectable between different human H2 influenza virus isolates from 1957 that were sensitive to inhibition by horse serum, indicating that minor antigenic variation occurs within the first year of appearance of the new subtype. Minor antigenic variation continued in the H2 viruses until 1961, but by 1962 antigenically distinguishable variants that could be discriminated with both monoclonal antibodies and postinfection ferret antisera predominated. Analysis of avian H2 influenza viruses with a panel of monoclonal antibodies indicated that antigenic variation occurs and that multiple different variants cocirculate in the population. There was no progressive antigenic change in the avian H2 influenza viruses with time, as was found with the human H2N2 strains. Topographical mapping of the H2 hemagglutinin by selection of antigenic variants with monoclonal antibodies and analysis of their reactivity patterns by HI showed overlap between the epitopes examined. These results may reflect restriction in the antibody repertoire of the mice used in preparation of the monoclonal antibodies or that the H2 hemagglutinin does not have such discrete nonoverlapping antigenic regions found in the early H3 influenza virus.

Antibodies, Monoclonal↗
PubMed1984

Antigenic and biological characterization of influenza virus neuraminidase (N2) with monoclonal antibodies.

Competitive radioimmunoassays using monoclonal antibodies established that the neuraminidase of A/RI/5+/57 (H2N2) influenza can be divided into four overlapping antigenic regions. Antigenic regions 1 and 4 are sufficiently far apart so that there was no competition between antibodies for these sites. Region 1 is conserved in neuraminidases from N2 viruses over a 10-year period, while the other regions changed antigenically during this time. The antibodies belonging to groups 2 and 3 completely inhibited catalytic activity on fetuin substrate, whereas antibodies in groups 1 and 4 inhibited weakly or not at all. Antigenic region 2 can be further divided into four overlapping areas (2a, 2b, 2c, and 2d) based on the reactivity patterns of monoclonal antibodies with antigenic variants, chemically modified neuraminidase, and the ability of the antibodies to inhibit enzyme activity of different molecular weight substrates. Previous studies [R. G. Webster, V. S. Hinshaw , and W. G. Laver (1982) Virology 117, 93-104; D. C. Jackson and R. G. Webster (1982) Virology 123, 69-77] characterized only region 2 of the neuraminidase molecule. Each of the monoclonal antibodies inhibited virus release from MDCK cells when incorporated in an agar overlay, and some antibodies in each group inhibited hemagglutination by intact virus, but only antibodies in group 2 neutralized virus in embryonated eggs and permitted selection of antigenic variants. The results indicate that antibodies to some antigenic sites on the neuraminidase may inhibit virus release more efficiently than others, depending on their relation to the enzyme active center. None of the monoclonal antibodies inhibited the hemolytic activity of viruses possessing N2. Based on antigenic mapping and biological properties of the monoclonal antibodies, a topographical map of the neuraminidase can be constructed. It is proposed that antigenic regions 1 and 4 are spacially separated and, based on their failure to inhibit biological activity, may be located on the bottom surface of the molecule; region 3 may be on the top surface of the molecule but at some distance from the catalytic center. Antigenic region 2 probably encompasses most of the top surface of the molecule; region 2d being closest to the enzyme center, with subregions 2a and 2b adjacent to it on the top surface. Chemical treatment of the neuraminidase with trinitrobenzenesulfonic acid (TNBS) causes modification of the 2b region, confirming the antigenic mapping results.

Animals↗
PubMed1984

Adrenal medulla imaging agents: a structure-distribution relationship study of radiolabeled aralkylguanidines.

Fourteen 125I-labeled aralkylguanidines were synthesized and evaluated as potential imaging agents for the adrenal medullae and tumors of adrenomedullary origin. These guanidines are radiotracer analogues of guanethidine, an antihypertensive agent thought to mediate neuron blockade by uptake into adrenergic nerves. Dog adrenal medullae were used as a model to test radiotracer affinity for catecholamine storage tissue. Tissue distribution studies revealed that a number of radioiodinated guanidines showed pronounced localization in the adrenal medullae following intravenous injection, in certain cases exceeding that of either (-)-[3H]norepinephrine or [14C]guanethidine. (m-[125I]Iodobenzyl)guanidine (m-IBG, 2b) gave the best combination of high concentration and selectivity. The low adrenomedullary affinity observed with [14C]guanidine and m-[125I]iodobenzylamine demonstrates the uniqueness of the aralkylguanidine structure. Preliminary evidence suggests that 2b is a storage analogue of norepinephrine. [125I]2a is now being used clinically in imaging and radiotherapy of catecholamine tumors, such as pheochromocytoma.

3-Iodobenzylguanidine↗
PubMed1984

Selection of monoclonal antibody to hCG which localizes in human choriocarcinoma growing in the Syrian hamster cheek pouch.

Six different monoclonal antibodies (MAbs) to human chorionic gonadotropin (hCG) were evaluated for radioimmunodetection in Golden Syrian Hamsters bearing a human choriocarcinoma in the cheek pouch (188-516 mIU of hCG/g). The affinity of three antibodies when determined were high and five of the antibodies were of IgG1 subclass. After i.v. injection of Staphylococcus Protein A (SPA) affinity purified radioiodinated antibodies, tissue distribution studies demonstrated that antibody 061 had higher target-to-nontarget ratios than the other five antibodies. The mean tumor:muscle ratio of 131I-MAb 061 was 27.5 in four animals, where the mean tumor:muscle ratio of four animals injected with 131I-labeled control monoclonal IgG1 Kappa from MOPC-21 murine myeloma (MOPC-21) was only 2.2. Sharp images of the cheek pouch tumor were obtained at five days following a tracer dose of 131I-061 MAb, but not with the same dose of a 131I-MOPC-21.

Animals↗
PubMed1984

Radioiodinated branched-chain fatty acids: substrates for beta oxidation? Concise communication.

Branched-chain iodinated fatty acids have been proposed for use as myocardial imaging agents. Several omega- iodoalkyl and omega- iodoaryl beta-methyl branched fatty acids have been synthesized and tested in rats. Myocardial activity levels at t = 5 min are affected by chain length for both alkyl and aryl acids, with chain lengths of 16 carbons possessing higher levels of activity than shorter lengths. Branching significantly lowers heart-to-blood ratios relative to straight-chain analogs. The degree of branching also affects radioactivity levels. Monoalkylation at the beta-carbon does not reduce the levels for omega- iodoalkyl fatty acids, but dialkylation reduces the levels significantly. Branching in the omega- iodoaryl series of fatty acids altered the time course of activity in the myocardium from a level of activity decreasing with time for the straight-chain acid to an essentially constant level of activity for the branched analogs.

Animals↗
PubMed1983

Antigenic variation in the influenza A virus nonstructural protein, NS1.

The antigenic structure of the nonstructural (NS1) protein encoded by influenza type A virus was examined using monoclonal antibodies prepared against purified NS1 inclusions isolated from the cytoplasm of infected cells. Topographical analysis by competitive radioimmunoassay indicated that three different overlapping antigenic regions were present on the NS1 of A/WSN/33 (H1N1). Immunoprecipitation studies using infected cell lysates showed that antigenic determinants on A/WSN/33 NS1 are common to NS1 proteins encoded by a wide range of viruses of human, swine, equine, and avian origin. Several avian strains, however, were found to encode antigenically variant NS1 proteins which had either extensive changes in one or more antigenic regions or small changes in epitopes within a region suggestive of antigenic drift. There was no correlation between surface antigen subtype and the antigenic profile of the NS1 protein. The antigenic relationships of NS1 proteins shown in this study are in agreement with the available sequence data.

Animals↗
PubMed1983

Changes in the antigenicity of the hemagglutinin molecule of H3 influenza virus at acidic pH.

In order to determine the location and biological significance of the acid-induced conformational change in influenza virus hemagglutinin (HA) reported by Skehel et al., monoclonal antibodies were prepared to the molecule before and after treatment at pH 5.0. These antibodies together with monoclonal antibodies to the different antigenic regions of the H3 HA were used in immunoprecipitation and ELISA binding studies to show that antigenic changes accompanied the conformational change in the HA. Treatment at pH 5.2 or less exposed new determinants on the HA while two antigenic regions, located at the tip and interface of the molecule at neutral pH, were lost or modified. Antigenic sites in the loop and hinge regions defined by the available monoclonal antibodies were not affected by the conformational change. Monoclonal antibodies specific for the acid-induced conformation efficiently inhibited hemagglutination of the virus at low pH but were extremely poor inhibitors of virus-induced red blood cell hemolysis at its pH optimum of 5.1. These antibodies were unable to neutralize viral infectivity under neutral or acidic conditions. Antibodies specific for the non-acid-treated HA conformation failed to inhibit hemagglutination at low pH values but were able to both inhibit hemolysis of red blood cells and neutralize virus infectivity. Residual unmodified HA after pH 5.0 treatment could explain the inhibition of hemolysis and infectivity by monoclonal antibodies in each of the different antigenic areas.

Animals↗
PubMed1983

Kinetics of tissue distribution and elimination of 4,4'-methylene bis(2-chloroaniline) in rats.

The tissue distribution kinetics and elimination of 4,4'-methylene bis(2-chloroaniline) (MBOCA) in rats was studied after a single dose of [14C]MBOCA (0.49 mg/kg body weight, i.v.). The highest concentrations of radioactivity were in the small intestine, liver, adipose, lung, kidney, skin, and adrenals. For most tissues, a rapid decrease in radioactivity was followed by a slower decrease except for the small intestine, adipose and skin which demonstrated transient increases. Subcellular distribution in liver at 1 h showed radioactivity in all cell fractions. Although very lipophilic, [14C]MBOCA was completely eliminated within 48 h with the major route via the feces (73.4%).

Adipose Tissue↗
PubMed1983

Heterogeneity of influenza B viruses.

Antigenic analysis of influenza B strains isolated in 1981-82 from England, Japan, and the USA, using a panel of monoclonal antibodies to the haemagglutinin of B/Oregon/5/80, showed considerable heterogeneity among the isolates, the majority of which had distinct reactivity patterns. Antigenically similar viruses were isolated from England, Japan, and the USA, and heterogeneity was detected among isolates from each country. Further studies are needed to determine whether this marked heterogeneity reflects different cocirculating strains or antigenic drift during an epidemic. The monoclonal antibodies failed to detect any difference between influenza B isolates from patients with Reye's syndrome and those circulating in the community.The monoclonal antibodies, each recognizing different determinants on the haemagglutinin of B/Oregon/5/80, were pooled and compared with ferret antisera to determine if the pool could be used as a reference reagent. The monoclonal antibody pool discriminated between isolates, contained no non-specific inhibitors of haemagglutination, and avoided the problems associated with differences between ferret antisera. In general, viruses that were shown to be antigenically different by ferret antisera were also different with the pooled monoclonal antibodies, but further studies are required to determine the optimal mixture of antibodies that will detect epidemiologically significant differences between strains.

Animals↗
PubMed1982

Antigenic determinants of influenza virus hemagglutinin--IX. The carbohydrate side chains from an Asian strain.

The heavy chain (HA1) of hemagglutinin from the Asian variant A/Japan/305/57 was examined to establish which of the four oligosaccharide units on this polypeptide could cross-react with host antigen. Cyanogen bromide and enzymic digests were fractionated and the resulting peptides screened for their ability to bind to antibodies raised against uninfected chick chorioallantoic membranes. This 324 residue HA1 polypeptide from the Asian strain A/Japan/305/57 is known to contain oligosaccharide units at asparagine residues 11, 23, 285 and at either 164, 165 or both. The peptides carrying the four carbohydrate units have been isolated, the monosaccharide compositions determined and the location of the fourth oligosaccharide shown to be Asn164 by direct amino acid sequence analysis. All four sugar units on A/Japan/305/57 HA1 are of the N-acetyllactosamine type, but only the one attached to Asn11 could cross-react with anti-(host antigen) antibodies. This antigenic carbohydrate unit on the Asian hemagglutinin heavy chain is located in a similar region to the two antigenically active oligosaccharides located on the hemagglutinin from the Hong Kong variant A/Memphis/102/72. The study was therefore extended to include the early Hong Kong variant A/Aichi/2/68, which has the same oligosaccharide distribution as A/Memphis/102/72 but different sugar compositions for the individual oligosaccharide units Like A/Memphis/102/72, only the N-acetyllactosamine units attached at positions 8 and 22 of A/Aichi/2/68 were capable of binding to anti-(host antigen) IgG. Finally, the light chain (HA2) polypeptides from A/Bel/42 (HO), the A/Japan/305/57 (H2) and the two Hong Kong variants A/Aichi/2/68 (X-31) and A/Memphis/102/72 were also examined and found to contain carbohydrate which is antigenically related to chick embryo host antigen.

Carbohydrates↗