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Biomedical subjects

B E Clarke

Publications and source records attributed to B E Clarke.

51 records · Page 3Linked to original sources
PubMed1987

A synthetic peptide which elicits neutralizing antibody against human rhinovirus type 2.

Synthetic peptides corresponding to six predicted immunogenic sites on human rhinovirus type 2 (HRV2) have been tested for their reactivity with an anti-virion antibody and for their ability to elicit neutralizing antibody. Four of the peptides reacted with HRV2 antiserum in an indirect ELISA. Rabbit antisera produced to three of these four peptides, one each from VP1, VP2 and VP3, reacted with the virus in an indirect ELISA and with the corresponding proteins by Western blotting. Furthermore, antiserum to one of the peptides, designed to cover the neutralization epitope NIm-II on VP2, not only reacted well in a sandwich ELISA and in an immunoprecipitation test but also neutralized virus infectivity.

Animals↗
PubMed1985

The sequence of foot-and-mouth disease virus RNA to the 5' side of the poly(C) tract.

The nucleotide sequence of foot-and-mouth disease virus (FMDV) RNA to the 5' side of the poly(C) tract (S fragment) has been determined for representatives of the A and O serotypes of the virus. The two S fragments differ in length by five nucleotides (nt), with 367 nt for O1 compared with 362 nt for A10, due to a number of insertions/deletions. However, the two sequences show 86% homology. There are no conserved open reading frames (ORFs). Secondary structure predictions reveal a high degree of potential base-pairing, such that the entire S fragment sequence can be folded into a hairpin structure.

Aphthovirus↗
PubMed1985

Benign fibrous histiocytoma of bone.

The clinical, radiologic, and pathological features of eight cases of fibrohistiocytic bone lesions histologically identical to the nonossifying fibroma of childhood are presented. They differed from the childhood lesion in their clinical and radiological features. They occurred in adults, and were frequently associated with pain in the absence of complicating fracture. They were not confined to the metaphysis of long bones. When metaphyseal, the lesions also frequently showed a tendency to involve the epiphysis. Others occurred in the diaphysis of long bones, in the pelvis, and in a rib. Three recurred locally, but none has metastasized. Other fibrohistiocytic and fibroblastic tumours of bone, including malignant fibrous histiocytoma, giant cell tumour, fibrosarcoma, and desmoplastic fibroma can be differentiated on radiological and histological features, and hyperparathyroidism may need to be excluded by biochemical investigations.

Adult↗
PubMed1985

Two initiation sites for foot-and-mouth disease virus polyprotein in vivo.

Typically, the translation of eukaryotic mRNAs into protein is initiated at a single site. However, we have recently shown that not one but two primary products, P20a and P16, are translated from the 5' end of the coding region of the genome of foot-and-mouth disease virus (FMDV). In this paper we show by partial protease digestion of these proteins that they differ only at their N termini, thus confirming the presence of two initiation sites for translation of FMDV RNA. Sequence analysis of two subtypes of the virus (A10 and A12) confirms the presence of two initiator AUG codons in the expected position on the genome. By correlation with protein synthesis data from these subtypes it appears that the relative use of each initiation site is dependent on its surrounding nucleotide sequence. In addition, the ratio of the two proteins when synthesized in vitro differs markedly from that when they are synthesized in vivo, suggesting the presence of a control mechanism for synthesis of P20a in vivo which may be absent in vitro. We also show that the cleavage site between these two proteins and the structural protein precursor, P88, is located closer to the N terminus of the polyprotein than has previously been reported.

Aphthovirus↗
PubMed1984

The complete nucleotide sequence of the RNA coding for the primary translation product of foot and mouth disease virus.

The complete nucleotide sequence of the coding region of foot and mouth disease virus RNA (strain A1061) is presented. The sequence extends from the primary initiation site, approximately 1200 nucleotide from the 5' end of the genome, in an open translational reading frame of 6,999 nucleotides to a termination codon 93 nucleotides from the 3' terminal poly (A). Available amino acid sequence data correlates with that predicted from the nucleotide sequence. The amino acid sequence around cleavage sites in the polyprotein shows no consistency, although a number of the virus-coded protease cleavage sites are between glutamate and glycine residues.

Amino Acid Sequence↗
PubMed1984

A new and improved method for 3'-end labelling DNA using [alpha-32P]ddATP.

We have synthesised dideoxyadenosine-5'-[alpha-32P]triphosphate [( alpha-32P] ddATP ) at a specific activity of 3000 Ci/mmol and directly compared it with cordycepin-5'-[alpha-32P]triphosphate [( alpha-32P] KTP ) as a means to 3'-end label DNA. The [alpha-32P] ddATP was found to be three to five times more efficient than [alpha-32P] KTP . Blunt and 3'-protruding ends were labelled more efficiently with [alpha-32P] ddATP using terminal transferase than were the 5'-ends with [gamma-32P]ATP using polynucleotide kinase by standard methods. This improvement in efficiency of labelling DNA and the simplicity of the method allows 3'-end labelling of DNA to become a realistic alternative to 5'-end labelling. We have also compared [alpha-32P] ddATP - and [alpha-32P] KTP -labelled DNA in Maxam and Gilbert sequencing procedures and find that both give equally good results.

Base Sequence↗
PubMed1984

Multiple proteases in foot-and-mouth disease virus replication.

Translation of foot-and-mouth disease virus RNA in a rabbit reticulocyte lysate for short time intervals resulted in the production of the peptides P20a , P16, and P88 (Lab, Lb, and P1) (R. R. Rueckert , Recommendations of the 3rd European Study Group on Molecular Biology of Picornavirus, Urbino , Italy, 1983). If further translation was prevented, the structural protein precursor P88 was not cleaved, even after prolonged incubation. This result indicates that the mechanism of the cleavage between P20a -P16 and P88 and of that between P88 and P52 (P2) differs from the mechanism of the secondary cleavages which produce the structural proteins. Furthermore, treatment of foot-and-mouth disease virus-infected cells with the protease inhibitor D-valyl phenylalanyl lysyl chloromethyl ketone prevented the in vivo cleavage between P20a -P16 and P88 but had no effect on any of the other cleavage events. These results suggest that the cleavage of the foot-and-mouth disease virus polyprotein utilizes two different host proteases.

Animals↗
PubMed1983

Synthetic peptides mimic subtype specificity of foot-and-mouth disease virus.

The major immunogen of foot-and-mouth disease virus (FMDV) is located between amino acids 141-160 of the capsid protein VP1. Synthetic peptides corresponding to the major immunogenic region give good neutralising antibody responses and protection in guinea pigs. To define more precisely the immunogenic site of the virus, we have examined serological differences between subtypes of the A serotype using synthetic peptides covering the 141-160 region. We show that these synthetic peptides carry determinants which mimic the subtype specificity of the virus. The correlation between these results and predictive structural models, based on the amino acid sequence, is discussed.

Amino Acid Sequence↗
PubMed1983

Origin of glomerular crescents in rabbit nephrotoxic nephritis.

The origin of glomerular crescents was investigated in an accelerated model of rabbit nephrotoxic nephritis. In rabbits immunised against sheep gamma-globulin, the administration of sheep nephrotoxic serum provoked cellular crescent formation affecting 30 to 90 per cent. of glomeruli at 6 days. The crescents were composed predominantly of cells with ultrastructural features of macrophages. In animals depleted of circulating leukocytes by whole body irradiation, with or without shielding of the kidney, crescent formation was inhibited despite severe glomerular damage and fibrin deposition in Bowman's space. These findings support the hypothesis that glomerular crescents develop principally from the emigration and accumulation of bone marrow-derived mononuclear cells.

Animals↗
PubMed1983

Macrophages and glomerular crescent formation. Studies with rat nephrotoxic nephritis.

A model of crescentic glomerulonephritis in the rat was developed, based upon an augmented form of nephrotoxic nephritis. The glomerular lesions were relatively mild, permitting an analysis of the morphologic events in crescent formation to an extent not possible in models in other species in which the sequence of changes is obscured by the severity of the inflammatory process. Monocytes and macrophages accumulated in glomerular capillary lumens and walls. Crescents were composed of cells with ultrastructural features indistinguishable from those of the intracapillary mononuclear cells. The findings support the view that the crescent cells are predominantly composed of macrophages derived from circulating monocytes.

Animals↗
PubMed1983

Gelatinous transformation of the bone marrow.

A case of gross macroscopic gelatinous transformation of bone marrow associated with prolonged malabsorption due to celiac disease is reported. Histochemical studies support the view that the extracellular myxoid tissue is composed predominantly of acid mucopolysaccharides, and electron microscopy shows extracellular fine fibrillar and granular material. The replacement of marrow in this condition is histochemically different from serous fat atrophy occurring in epicardial fat, and it is suggested that it results from an increase in normal ground substance. It must be differentiated from metastatic signet ring adenocarcinoma.

Adenocarcinoma, Papillary↗
PubMedDate not supplied

Synthesis and secretion of a functional antibody in a vaccinia virus expression system.

A humanized rat monoclonal antibody (Campath 1H) has been expressed in HeLa cells using recombinant vaccinia viruses. Heavy and light chain recombinant viruses were constructed separately and when grown independently produced proteins of the expected molecular weights. Expressed heavy chain was entirely intracellular but light chain was mainly excreted and processed. When cells were infected at high multiplicity with both heavy and light chain recombinants a proportion of the heavy chain was then found in the extracellular medium. This secreted heavy chain was shown to be associated with light chain as judged by co-electrophoresis in non-reducing SDS polyacrylamide gels and by co-purification on protein-A sepharose. The secreted heavy and light chain complexes were functionally active as an antibody, with activity comparable to authentic Campath 1H antibody as assessed by ELISA, T-cell binding and antigen binding assays. Production of antibody in this system was achieved in the absence of serum, which is an important consideration in the production of monoclonal antibodies (MAbs). The amount of antibody produced was 0.2-0.4 micrograms/10(6) cells without optimization of expression levels. The wide host cell range of vaccinia virus together with the recently developed methods for increasing expression levels make this an attractive candidate as a flexible general vehicle for producing MAbs.

Animals↗
PubMedDate not supplied

Chemical basis of antigenic variation in foot-and-mouth disease virus.

One of the difficulties in controlling foot and mouth disease by vaccination is the occurrence of the virus as seven distinct serotypes because immunity conferred by vaccination against one serotype leaves the animals susceptible to infection by the other six. Moreover, the antigenic variation, even within a serotype, can be so great that immunity against the homologous strain of virus need not necessarily ensure protection against infection by other viruses within that serotype. Here we report the separation of three natural antigenic variants, distinguishable in cross-neutralization tests from an isolate of foot-and-mouth disease virus (FMDV). The serological differences could also be demonstrated by antisera elicited by synthetic peptides corresponding to residues 141-160 of the capsid polypeptide VP1, showing that this region contains a major immunogenic site of the virus. The results have practical implications for the choice of viruses for vaccine production.

Amino Acid Sequence↗
PubMedDate not supplied

Improved immunogenicity of a peptide epitope after fusion to hepatitis B core protein.

Synthetic vaccines for viral diseases can use defined regions of viral proteins as immunogens: the peptide sequence of amino acids 141-160 of the VP1 protein of foot and mouth disease virus (FMDV) elicits virus-neutralizing antibodies to protect guinea pigs, cattle and pigs either when coupled to a carrier protein or when administered in liposomes or in incomplete Freund's adjuvant. The immune response to these peptides is much lower than that to complete virus particles and the same sequence fused to the N terminus of beta-galactosidase did not produce a more potent immunogen than synthetic peptide alone. We report here an expression system for immunogenic epitopes linked to a carrier protein, hepatitis B core antigen, to form part of a virus-like complex which can present these epitopes to the immune system at high density. The immunogenicity of these structures approaches that of FMDV particles.

Animals↗