Biomedical subjects

A Berge

Publications and source records attributed to A Berge.

18 recordsLinked to original sources

PubMed1998

Identification of an insertion sequence located in a region encoding virulence factors of Streptococcus pyogenes.

An insertion sequence, IS1562, was identified in a Streptococcus pyogenes strain of the clinically important M1 serotype. IS1562 is located in the mga regulon between the genes coding for the M protein and the C5a peptidase, both important virulence factors. The same or similar insertion sequences were found in most S. pyogenes strains, but the chromosomal location differed among isolates.

Adhesins, Bacterial↗

PubMed1997

Streptococcal protein H forms soluble complement-activating complexes with IgG, but inhibits complement activation by IgG-coated targets.

Protein H, a surface protein of Streptococcus pyogenes interacting with the constant Fc region of IgG, is known to be released from the streptococcal surface by a cysteine proteinase produced by the bacteria. Poststreptococcal glomerulonephritis and rheumatic fever are conditions in which immune complexes and autoimmune mechanisms have been suggested to play pathogenetic roles. The present study demonstrates that addition of protein H to human serum produces complement activation with dose-dependent cleavage of C3. The activation was IgG-dependent and the result of complexes formed between IgG and protein H. These complexes were size heterogeneous with molecular masses of 400 kDa to 1.4 MDa. Using complement-depleted serum reconstituted with complement proteins, the activation by protein H was found to be dependent of the classical, but independent of the alternative pathway of complement. In contrast to results of experiments based on soluble protein H.IgG complexes, complement activation was inhibited by protein H when IgG was immobilized on a surface. The interaction between C1q and immunoglobulins represents the first step in the activation of the classical pathway, and protein H efficiently inhibited the binding of C1q to IgG immobilized on polyacrylamide beads. Protein H reduced C3 deposition on the IgG-coated beads and inhibited immune hemolysis of IgG-sensitized erythrocytes. Finally, significantly less C3 was deposited on the surface of protein H-expressing wild-type streptococci than on the surface of isogenic mutant bacteria devoid of protein H. The results demonstrate that protein H.IgG complexes released from the streptococcal surface can produce complement breakdown at the sites of infection, whereas complement activation on bacterial surfaces is inhibited. This should have important implications for host-parasite relationships. In addition, soluble protein H.IgG complexes might contribute to immunological complications of streptococcal infections.

Antigens, Bacterial↗

PubMed1997

Biological effects of the immunomodulator beta 1-3D polyglucose are strongly potentiated by conjugation to biodegradable microbeads.

It is demonstrated that the biological effects of the immunomodulator beta 1-3D polyglucose, when covalently linked to polymethacrylate or biodegradable albumin microbeads, are strongly potentiated. The potentiation is recorded as an increased protection effect of the conjugates in Escherichia coli sepsis in mice, and as increased IL-1 production by murine macrophages in vitro.

Adjuvants, Immunologic↗

PubMed1995

Streptococcal cysteine proteinase releases biologically active fragments of streptococcal surface proteins.

Streptococcus pyogenes are important pathogenic bacteria which produce an extracellular cysteine proteinase contributing to their virulence and pathogenicity. S. pyogenes also express surface molecules, M proteins, that are major virulence determinants due to their antiphagocytic property. In the present work live S. pyogenes bacteria of the M1 serotype were incubated with purified cysteine proteinase. Several peptides were solubilized, and analysis of their protein-binding properties and amino acid sequences revealed two internal fibrinogen-binding fragments of M1 protein (17 and 21 kDa, respectively), and a 36-kDa IgG-binding NH2-terminal fragment of protein H, an IgGFc-binding surface molecule. M protein also plays a role in streptococcal adherence, and removal of this and other surface proteins could promote bacterial dissemination, whereas the generation of soluble complexes between immunoglobulins and immunoglobulin-binding streptococcal surface proteins could be an etiological factor in the development of glomerulonephritis and rheumatic fever. Thus, in these serious complications to S. pyogenes infections immune complexes are found in affected organs. The cysteine proteinase also solubilized a 116-kDa internal fragment of C5a peptidase, another streptococcal surface protein. Activation of the complement system generates C5a, a peptide stimulating leukocyte chemotaxis. C5a-mediated granulocyte migration was blocked by the 116-kDa fragment. This mechanism, by which phagocytes could be prevented from reaching the site of infection, may also contribute to the pathogenicity and virulence of S. pyogenes.

Adhesins, Bacterial↗

PubMed1993

PAM, a novel plasminogen-binding protein from Streptococcus pyogenes.

The ability of group A streptococci to bind human plasminogen and plasmin has attracted interest, because it could provide the bacteria with a mechanism for invasion. M or M-like proteins account for the binding of several plasma proteins to group A streptococci. To investigate whether M or M-like proteins were responsible for the binding of plasminogen to group A streptococci, acid-extracted material from a type M53 streptococcal isolate was tested for its ability to bind plasminogen. Indeed, a 42-kDa plasminogen-binding protein was solubilized. Two oligonucleotides homologous with conserved sequences in known M protein genes were used as primers in the polymerase chain reaction, with chromosomal DNA from the M53 isolate. When cloned and expressed in Escherichia coli, a resulting fragment encoded a 43-kDa plasminogen-binding protein. Nucleotide sequence determination of the gene fragment revealed an open reading frame encoding a polypeptide of 43,580 Da, which matched the amino-terminal amino acid sequence of the plasminogen-binding protein extracted from M53 streptococci. The DNA sequence data also proved the relationship of the encoded protein, named PAM, to the M proteins. The plasminogen-binding domain was mapped to the amino-terminal third of PAM. Plasminogen absorbed by M53 streptococci or by immobilized PAM could be activated by streptokinase. The results provide further evidence of the diversity of the M protein family and suggest a new mechanism whereby these proteins contribute to the virulence of group A streptococci.

Amino Acid Sequence↗

PubMed1993

Monodisperse magnetic polymer particles. New biochemical and biomedical applications.

The method of activated swelling of polymer particles developed by the authors allows the preparation of monodisperse spherical beads of predictable size from 1 to 100 microns in diameter. The polymer particles may be prepared from a number of different monomeric materials and with various morphologies including macroporous structures. The porous beads form the basis for magnetizable monodisperse polymer particles which have magnetic iron oxides distributed as small grains all through the volume of the beads. The magnetic particles are being used extensively for selective cell separation and for immunomagnetic separation within microbiology and molecular biology. A review of recent work within these fields is given. New methods for positive cell separation are announced.

Bone Marrow Purging↗

PubMed1993

Homogeneous immunofluorometric assays of alpha-fetoprotein with macroporous, monosized particles and flow cytometry.

We evaluated two homogeneous immunofluorometric assays (IFMAs) of alpha-fetoprotein (AFP) based on new macroporous acrylate particles combined with flow cytometry. The standard IFMA, requiring 1 h of incubation, provided a working range from 1.8 to > 900 kIU/L (CV < 10%) and a detection limit of 0.6 kIU/L. Use of overnight incubation and a lower particle concentration extended the working range by 1 decade in the lower end. Analytical recoveries for the standard IFMA varied between 97% and 108%. The slope and y-intercept of the regression line correlating measurements by the standard IFMA and a routine immunoradiometric assay were not significantly different from 1 and 0, respectively (P > 0.5), and the correlation coefficient was 0.996. High precision and warning of spuriously high measurements were obtained by including in each sample separate particle types for detecting instrument instability and measuring nonspecific binding only.

Acrylates↗

PubMed1988

Monosized, magnetic polymer particles: their use in separation of cells and subcellular components, and in the study of lymphocyte function in vitro.

By employing the principles of "activated swelling", monosized, superparamagnetic polymer particles have been prepared ranging in size from 1-100 microns. Both during and after the swelling process, the particles can be modified to meet a series of specific demands making them potentially very interesting for many separation and assay purposes. Using monoclonal antibodies to direct the magnetic beads to their targets, immunomagnetic separation has turned out to be one of the most specific, reliable and, above all, the fastest technique available today to isolate particulate material for further studies. So far, most efforts have been concentrated on methodology for fractionation of cells in suspension, such as removal of tumour cells from bone marrow or isolation of lymphoid cells from peripheral blood. These studies have both established the parameters necessary for optimal performance and at the same time laid the groundwork for future developments making immunomagnetic separation an exciting new tool in many research areas. High speed and specificity are the most conspicuous features of immunomagnetic cell separation. These properties have been exploited in the successful development of a new technique for tissue typing of cells directly from peripheral blood specimens. Both higher sensitivity and specificity have been obtained. The same principles can be used for fast and safe quantification of cell populations and subpopulations in blood and cell suspensions. The functions of, and interactions between, peripheral blood cell populations or subpopulations in the immune response have also been studied with high precision. The significance of direct cell contact on the one hand, and soluble factors on the other, can now be established in detail. Immunomagnetic beads have also been used to study the interaction between various T lymphocyte membrane molecules in the early phases of the activation process. Finally, the usefulness of specially developed particles for the fractionation of subcellular components is described.

Antibodies, Monoclonal↗

PubMed1985

In vivo activation of mouse macrophages with beta-1,3-D-glucan-derivatized plastic beads.

Macrophages obtained from animals treated with beta-1,3-D-glucan-derivatized plastic beads were greatly stimulated, as judged by morphology, esterase release, and cytostatic effect on L-929 tumour cells in vitro. The pretreatment of mice with such beads conferred an apparent absolute local resistance to an otherwise lethal pneumococcal infection but had no effect on the growth of intraperitoneal AA ascites sarcoma. Moreover, peritoneal cells from animals pretreated with glucan beads did not protect the animals in a Winn assay.

Animals↗

PubMed1984

Hydrophilic monodisperse particles as solid-phase material in immunoassays: comparison of shell-and-core particles with compact particles.

Hydrophilic monodisperse shell-and-core particles with a density of 1.07 were superior to heavier compact particles as a solid-phase material for immunoassays. The shell-and-core particles formed a semistable suspension for 24 h and were easily collected by centrifugation. The hydroxyl groups of the particles were activated with two sulfonyl chlorides. The most reactive one, tresyl chloride, gave rapid chemical coupling of antibodies, whereas tosyl chloride favored a rapid hydrophobic adsorption which was followed by slow chemical coupling. The solid-phase sheep antirabbit IgG made was used as a separation agent in several immunoassays and gave solid-phase primary antibodies by immunoadsorption of rabbit antibodies.

Animals↗

PubMed1983

Excess antibody immunoassay for rat glandular kallikrein. Monosized polymer particles as the preferred solid phase material.

The development of an excess antibody assay for rat glandular kallikrein is described. This assay permits immunological determination of kallikrein as well as a simultaneous specific measurement of kallikrein enzymatic activity. The assay is based on coupling of immunopurified anti-kallikrein immunoglobulin to a solid phase. In a first incubation step, kallikrein was bound to the immobilized antibody. Determination of kallikrein was subsequently done in a second incubation step; immunologically by addition of iodinated anti-kallikrein antibody, or enzymatically by a kallikrein substrate. Enzymatic quantification could also be followed by immunological measurements on the same sample. Comparison of Sepharose, cellulose, and acrylate based polymer particles proved the latter to be the best matrix in this assay. The main advantage of the polymer particles was the low non-specific binding of labelled antibody.

Animals↗

PubMed1976