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Biomedical subjects

J Sjöquist

Publications and source records attributed to J Sjöquist.

At least 73 records · Page 4Linked to original sources
PubMed1979

Effect of protein A of Staphylococcus aureus on the binding of monomeric and polymeric IgG to Fc receptor-bearing cells.

The effect of protein A of Staphylococcus aureus (SpA) on the binding of rabbit IgG to the Fc receptors of mouse lymphocytes and macrophages was found to correlate with the aggregation of the IgG ligands. After binding anti-erythrocyte IgG complexed with SpA, the cells were able to attach to and kill erythrocyte indicator cells with a higher efficiency than lymphoid cells treated with anti-erythrocyte IgG alone. The amount of anti-peroxidase IgG which can be bound to effector cells was not changed by reaction with SpA. In contrast, the binding to cells of IgG-coated erythrocytes and of anti-peroxidase IgG complexed with peroxidase was substantially reduced by reaction with SpA. The results are compatible with the presence of two distinct Fc receptors, one for cytophilic monomeric IgG and another for polymeric (antigen-complexed) IgG.

Animals↗
PubMed1978

II. Ultrastructural differences in attachment patterns of indicator cells to target cells in the rosette-forming process.

A comparison was made of the electron microscopic appearances of rosettes formed between mouse spleen lymphocytes and indicator cells with the ligands exposed on their surfaces either in clusters or in continuity. Sheep red blood cells (SRBC) coated with protein A of Staphylococcus aureus or human IgG by the chromium chloride technique were used as indicator cells with cluster distribution of the ligand. RBC coated with rabbit IgG anti-sheep red blood cells or staphylococci were used as indicator cells with uniform distribution of the ligand. Results show that the appearance of the zones of contact (point attachment or broad zone attachment) between lymphocytes and indicator cells are mainly influenced by the distribution of the specific ligands on the surface of the indicator cells and do not reflect the pattern of distribution of receptors on lymphocytes.

Animals↗
PubMed1978

Separation of cells by affinity chromatography on SpA-sepharose 6MB.

A cell separation technique was designed which was based on the reaction of protein A of Staphylococcus aureus (SpA) with the Fc regions of IgG antibodies. SpA covalently linked to Sepharose 6MB is able to fix antibody-bearing cells, while non-fixed cells are removed by washing. The fixed cells can be released by dog IgG or by mechanical treatment. As an example of the use of the technique, Ig-bearing cells were isolated from mouse spleen cells treated with rabbit IgG anti-mouse Ig antibody; the purity of the Ig-bearing lymphocytes so isolated was better than 90%. The viability and the ability of the cells to shed the antibodies from their surface were not significantly impaired by the fractionation method. The technique can be generally applied to cell separation providing IgG antibodies against specific surface markers are available.

Animals↗
PubMed1978

Crystallization, crystal structure analysis and atomic model of the complex formed by a human Fc fragment and fragment B of protein A from Staphylococcus aureus.

Crystals of the complex formed by human Fc fragment and fragment B (FB) of protein A from Staphylococcus aureus were prepared and the crystal structure determined at high resolution by multiple isomorphous replacement. Phase were improved considerably by combining these phases with calculated phases from the Fc component. FB is a small globular protein built of three parallel helices arranged in a triangular array. It binds by the first two helices of Fc and is attached to segments of CH2 and CH3. The CH3 module is unchanged between complex and Fc fragment crystals, but CH2 changes its position slightly relative to CH3. In addition, the upper third of CH2 is disordered in the complex crystals. Possible sources of this disorder are discussed.

Antigen-Antibody Complex↗
PubMed1978

Effect of cell density on the disappearance of cell surface immunoglobulins.

Mouse macrophages and human peripheral blood lymphocytes exhibiting on the membrane autologous IgG as well as mouse thymocytes coated with human IgG, showed a strong cell concentration dependence of surface IgG disappearance during an incubation at 37 degrees C. Mouse spleen lymphocytes treated with anti-mouse Ig behaved in the same way regarding the disappearance of surface immunoglobulins, while the redistribution of ligands (cap formation) was not dependent on cell concentration.

Animals↗
PubMed1976

Arming of lymphoid cells by IgG antibodies treated with protein A from Staphylococcus aureus.

Mouse spleen lymphocytes treated with rabbit IgG anti-sheep erythrocytes (SRBC) complexed with protein A of Staphylococcus aureus (SpA) form rosettes with SRBC. The attachment of SRBC to lymphocytes was due to the binding of the SpA-IgG antibody complex to the surface of the lymphocytes and was thus considered "arming" of the cells. Normal mouse spleen cells "armed" with SpA-rabbit IgG anti-chicken erythrocytes (CRBC) kill specifically 51Cr-labeled CRBC "in vitro" in the absence of free antibodies. The killing by these "armed" cells is an effect of the cell-bound SpA-IgG antibody complex. Both the SRBC rosette formation and the cell-mediated CRBC killing was dependent on the concentration of the SpA-IgG antibody complexes used for "arming" the cells. A 100-fold increase in rosette formation or in killing of target cells was recorded for lymphocytes treated with SpA-IgG antibody complexes in comparison with cells treated with noncomplexed IgG antibodies. The specific binding of SpA-IgG antibody complexes to the Fc receptors of mouse spleen cells was demonstrated by inhibition studies. More than 60% inhibition of the rosette formation and in the killing of target cells was shown for cells treated with normal rabbit IgG or its Fc fragment before addition of the SpA-IgG antibody complex.

Animals↗
PubMed1976

A method for the detection and quantitation of Fc receptor sites on cells.

A new rosette technique for identification of Fc-receptor-bearing cells is based on the ability of sheep erythrocytes coated with protein A of Staphylococcus aureus (ES) to form rosettes with cells treated with monomeric IgG or aggregated IgG. The IgG is attached to lymphocytes through its Fc region and binds to a trypsin-resistant but pronase-sensitive receptor (considered an Fc receptor). The ES rosette technique facilitates studies of the interaction of IgG with Fc receptor sites; the binding of any IgG preparation reacting with protein A of Staphylococcus aureus (SpA) can be studied by the technique. Inhibition of rosette formation by SpA was used for quantitation of IgG fixed to the cell surface (that is, the number of Fc receptors/cell). The sensitivity of the method permits quantitation of less than 10(5) IgG molecules bound to the Fc receptors. The relative affinity constant between Fc receptors and IgG ligands was estimated by plotting the percentage of ES rosettes as a function of IgG concentration and calculating the reciprocal of the IgG concentration giving half the maximal number of ES rosettes.

Animals↗
PubMed1975

Immunologically active and structurally similar fragments of protein A from Staphylococcus aureus.

To study the active site(s) in protein A, partial tryptic digestions of the protein and of intact Staphylococcus aureus were performed. Fragments which bind to the Fc-part of human IgG were isolated by affinity chromatography on IgG-Sepharose 4B and purified by ion-exchange chromatography on phosphocellulose. From a partial tryptic digest of pure protein A at 30 degrees C, pH 8.2 for 30 min we have isolated and characterized six active fragments with molecular weights ranging from 6000 to 8000. Two active fragments, obtained in high yields by digestion at pH 7.2 of intact protein-A-containing bacteria, were shown to be similar to two of the six characterized fragments from the digest of pure protein A. All fragments appeared to have similar amino acid sequences, judged by peptide mapping, specific staining and amino acid analysis; some are very possibly overlapping peptides. Each fragment probably contains only one active site region since all are monovalent in the Fc-reaction when studied with a hemagglutination technique. The maximal molar yield of active fragments obtained from the digestion of pure protein A accounts for about 210% of the amount of protein A used. Thus protein A, suggested to consist of repeating units, should exhibit at least three similar if not identical active regions.

Amino Acid Sequence↗
PubMed1975

A human lymphoid cell line with an IgG-like membrane component.

A human lymphoid cell ine (S95) with surface-located IgG is described. The line was derived from a patient with Hodgkin's disease and is aneuploid. Its characteristics are suggestive of a malignant B lymphocyte derivation, but no ultimate proof of malignancy can be forwarded. The line produces immunoglobulin (IgG) which appears to be devoid of light chain. The amount of IgG detectable at the cell surface by inhibition of the rosette formation by protein A-coated sheep red blood cells with protein A was 5-10 ng/10(6) cells corresponding to 70 000 - 140 000 molecules/cell. The immunoglobulin is membrane bound. Partial characterization of the IgG-like protein shows that it has a molecular weight of 50 000 Daltons and that the molecule lacks some of the normal gamma-chain determinants.

Cell Line↗
PubMed1975

Demonstration and assaying of IgG antibodies in tissues and on cells by labeled staphylococcal protein A.

Staphylococcal protein A (SpA) labeled with FITC was used to detect IgG antibodies to tissue and cell surface antigens. Radioiodinated SpA was employed in a radioimmuno technique to assay IgG antibodies which had reacted with cell membrane antigens. Red blood cells coated with SpA were used as an indicator system in the mixed haemadsorption technique to detect IgG antibodies on tissue culture cells. The results showed that SpA was highly specific in the various systems tested, although it was not as sensitive in the immunofluorescence test as FITC anti-IgG. It was confirmed that SpA reacts with IgG antibodies of human, monkey, rabbit, sheep and mice origin. The reactivity with human and monkey IgG was stronger than with IgG of the other species. We consider labeled SpA as an alternative to monospecific anti-IgG for detection and assaying of IgG antibodies to tissue and cell surface antigens.

Adrenal Glands↗
PubMed1975

Cell separation by staphylococcal protein A-coated erythrocytes.

A method for cell separation according to cell surface markers is described. Heat-aggregated human IgG or IgG antibodies, raised to surface markers of mouse lymphocytes-for example, Ig or theta antigen-are allowed to react with the lymphocytes. When these cells are incubated with sheep erythrocytes coated with protein A of Staphylococcus aureus, rosettes are formed. These rosettes can be separated from nonrosetted lymphocytes by density gradient centrifugation. The erythrocytes of the rosettes are disrupted by complement action or by osmotic shock, and the lymphocytes are recovered. Immunoglobulin-protein A complexes are shed off the cell surfaces by cultivation. In this way cells with, as well as those without, surface markers could be obtained with high purity (up to 80%) and with high viability.

Animals↗
PubMed1975

Separation of lymphocytes by specific adherence to cellular monolayers containing protein A of Staphylococcus aureus.

A technique was developed for the isolation of lymphocytes according to their surface antigenic markers. The method was based on the general reaction of protein A (SpA from S. aureus) with the Fc-part of an IgG antibody. Monolayers of S. aureus or Spa-coated sheep red blood cells (SpA-SRBC) fix antibody-charged cells specifically; non-fixed cells are easily removed by washing. Alternatively, the monolayers can be treated with a cell surface specific antibody prior to addition of non-charged cells. Monolayer-adhered cells were detached by the addition of lysostaphin (bacterial monolayer) or ammonium chloride (SpA-SRBC monolayer). As an example, Ig-bearing cells were isolated from mouse spleen lymphocytes using a specific rabbit anti-mouse Ig serum for charging either the cells or the monolayers. The recovery of Ig-bearing cells was approximately 84%. The purity of the cells was approximately 83% and the viability 89%. Antibody-SpA complexes on the surface of isolated cells were removed either by trypsin treatment or by cultivation. Ig on the cell surface is restored after 16 hr of cultivation.

Ammonium Chloride↗
PubMed1974

Density gradient separation of lymphoid cells adhering to protein-A-containing staphylococci.

A cell separation technique was designed based on the interaction between cell-surface-bound IgG and protein A of Staphylococcus aureus. The density of lymphoid cells coated with IgG antibodies against one of the surface markers was increased by adherence of staphylococci. Cells with adhering bacteria were separated from cells without bacteria by density gradient centrifugation in 11.5% sodium metrizoate. Bacteria were removed from the lymphoid cells by lysostaphin digestion. The purity of separated cells was approximately 95% even when the proportion of a specific cell population was below 10% in the initial mixture. The viability and the ability of cells to multiply in vitro were not significantly impaired by the fractionating procedure. The technique can generally be applied for cell separation, provided antibodies of the IgG class against specific surface markers are available.

Antigen-Antibody Reactions↗