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S Ohlson

Publications and source records attributed to S Ohlson.

13 recordsLinked to original sources
PubMed1989

Analysis of a glucose-containing tetrasaccharide by high-performance liquid affinity chromatography.

In the present work we have explored conditions for using a pulsed amperometric detector for on-line analysis of oligosaccharides eluted from a high-performance liquid affinity chromatography column. A monoclonal antibody that specifically binds a glucose-containing oligosaccharide is coupled to a SelectiSphere-10-activated tresyl column. The system is eluted isocratically and easily detects 10 ng of the oligosaccharide with a linear response up to 250 ng. Analysis of both serum and urine samples from normal individuals and patients with acute pancreatitis gives a single retarded peak with a retention time identical to that of authentic (Glc)4. Retarded material pooled from several analyses of urine was positively identified as (Glc)4 by GC-MS analysis. As this method requires little cleanup and no chemical derivitization of the sample and is performed rapidly (less than 20 min) at sensitivities of at least 10 micrograms/liter in biological fluids, it represents a substantial improvement over previous GC-MS, radioimmunoassay, and enzyme-linked immunoadsorbent assay methods used to determine (Glc)4.

Animals↗
PubMed1988

A novel approach to monoclonal antibody separation using high performance liquid affinity chromatography (HPLAC) with SelectiSpher-10 protein G.

Protein G, a bacterial cell wall protein extracted from strains of Streptococci, has been employed as a ligand in high performance liquid affinity chromatography (HPLAC) for separation of monoclonal antibodies. Examples are given of rapid high-resolution separations of rat and mouse monoclonal antibodies belonging to various subclasses. In comparison with protein A chromatography, we were able to show superior binding characteristics for SelectiSpher-10 protein G columns under conditions of 'low' ionic strength (about 0.1 M) and neutral pH (pH approximately 7). The monoclonal antibodies were isolated in high purity (greater than 90%) and with good recovery of specific activity (80-100%). We believe that the HPLAC technology based on SelectiSpher-10 protein G is of potential value in the analysis and purification of monoclonal antibodies from various species and subclasses.

Animals↗
PubMed1988

Novel approach to affinity chromatography using "weak" monoclonal antibodies.

Affinity purification generally relies on specific high-affinity recognition between two species of biological molecules: one molecular species (the ligate) dissolved in a mobile phase is selectively adsorbed to the other species (the ligand) coupled to a solid support. Desorption of the ligate often requires harsh conditions that degrade biological activity of the purified product. As an alternative to this general procedure, we have studied affinity chromatography in a weak affinity mode, where ligand-ligate interactions are in dynamic equilibrium. Ligates recognized with low affinities (dissociation constant greater than 10(-4) M) elute from affinity columns under mild, isocratic conditions as retarded peaks, separated from noninteracting solutes that elute in the void volume. To illustrate the procedure, we report chromatography of an oligosaccharide on a 2-ml column containing 86 mg of a monoclonal antibody coupled to 10-micron microparticulate silica particles. Using a temperature-sensitive antibody, we observed that when the ligand-ligate dissociation constant is greater than 10(-3) M, performance of the system exceeds 300 theoretical plates/10 cm column length and approaches the efficiencies generally associated with high-performance liquid chromatography.

Antibodies, Monoclonal↗
PubMed1988

Profiling of circulating immune complexes by high performance liquid affinity chromatography (HPLAC) on a protein A-silica matrix.

Protein A, a cell wall protein found in bacterial strains of Staphylococcus aureus, has been extensively used in the analysis and purification of immunoglobulins and immune complexes. By binding protein A to microparticulate silica (high performance liquid affinity chromatography, HPLAC), a rapid and efficient chromatographic system was obtained for the separation and analysis of circulating immune complexes. The method was applied to the separation of artificial immune complexes as well as to plasma samples from patients with immune complex associated diseases such as SLE and RA. It was possible to distinguish certain subpopulations of circulating immune complexes by performing pH gradients on the protein A silica HPLAC column.

Antigen-Antibody Complex↗
PubMed1988

High-performance liquid affinity chromatography: rapid immunoanalysis of transferrin in serum.

We describe a new method for quantitatively measuring substances of clinical interest by high-performance liquid affinity chromatography (HPLAC). As a model system we selected analysis for transferrin in human serum with immobilized antibodies in a high-performance liquid chromatographic system. SelectiSpher-10 Activated Tresyl columns (5 or 10 x 0.5 cm) were used for in situ coupling of polyclonal antibodies to transferrin. The amount of transferrin eluted was determined by integrating the eluted peak at 280 nm. The whole analytical procedure--including injection of sample, washing, elution, and analysis of data--takes only 7 min. We characterized the HPLAC system for analysis of transferrin in several ways: intra-assay CV approximately 3%; inter-assay CV 2-9%; linear response up to 1 mg/mL column volume; detection limit approximately 3 micrograms; analytical recovery 98% +/- 2%; purity of eluted sample greater than 95% (SDS-PAGE). The HPLAC method was compared with "rocket" immunoelectrophoresis, a commonly used method of analysis for transferrin, and there was excellent correlation between the two methods (r = 0.96, n = 60). Benefits of this HPLAC technique include high precision, rapid analysis, and simplified sample handling.

Chromatography, High Pressure Liquid↗
PubMed1988

The development of a C1q anti-C1q immunoadsorbent for removal of immune complexes from plasma.

An immunoadsorbent based on immobilized C1q has been developed to remove possibly pathogenic immune complexes from plasma deriving from patients suffering from systemic lupus erythematosus or rheumatoid arthritis. Traditional immobilization procedures based on, e.g., cyanogen bromide activation could not be used to produce an efficient adsorbent. However, by using antibodies directed towards C1q as handles for the immobilization of C1q it was possible to make an adsorbent that efficiently bound immune complexes in plasma. The capacity of the C1q anti-C1q adsorbent to bind artificial immune complexes such as aggregated human globulins or immune complexes from various plasma samples was evaluated. Both batch and column experiments were conducted. The typical capacity in batch was about 1 mg immune complexes/ml gel when incubated with patient plasma samples with high titers of immune complexes. Special attention has to be paid to leakage of undesirable components from the adsorbent. It was found that leakage of C1q occurred but it was not more than after covalent immobilization procedures such as cyanogen bromide.

Antigen-Antibody Complex↗
PubMed1987

High-performance liquid affinity chromatographic separation of mouse monoclonal antibodies with protein A silica.

Protein A, a bacterial cell wall protein found in Staphylococcus aureus, has been widely used for the analysis of immunoglobulins. By attaching protein A to a microparticulate silica support, a rapid and efficient chromatographic sorbent has been created for the separation of monoclonal antibodies. Examples are given of rapid separations (within 10 min) of murine monoclonal antibodies, belonging to various IgG subclasses and including IgG1. The monoclonal antibodies were isolated with a high purity and with 60-90% recovery of activity. The high-performance liquid affinity chromatography technique based on protein A provides a useful method for monitoring monoclonal antibodies in crude samples, such as ascites and cell culture supernatants.

Animals↗
PubMed1985

Detection of circulating immune complexes by PEG precipitation combined with ELISA.

An assay to detect circulating immune complexes is described and exemplified with serum from SLE and RA patients. The method is based on selective precipitation of immune complexes with PEG followed by a specific ELISA assay to detect the immunoglobulins present in the precipitate. The immunoglobulins of the precipitate were then monitored by their capacity to bind protein A and dDNA. With the PEG-protein A-ELISA procedure immune complexes were detected in 46% of the sera from SLE patients compared with 68% when using the PEG-DNA-ELISA.

Antigen-Antibody Complex↗