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Sökning: LAR1:liu > Linnéuniversitetet > Ohlson Sten

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  • Aldén, Anna, et al. (författare)
  • HPLC analysis of carbohydrate deficient transferrin isoforms isolated by the Axis-Shield %CDT method
  • 2005
  • Ingår i: Clinica Chimica Acta. - 0009-8981. ; 356:1-2, s. 143-146
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: Carbohydrate-deficient transferrin (CDT) is elevated during prolonged overconsumption of alcohol and CDT is considered to be the most specific biochemical marker for alcohol overconsumption. However, an accurate method for analysing CDT is necessary because the test is frequently used for example in legal matters. Methods: Patient serum samples were analysed with the Axis-Shield %CDT and eluates were pooled together. Transferrin was purified from the pool by affinity chromatography and further analysed with HPLC to determine the ratios of different transferrin isoforms. Results: In the eluates using the Axis-Shield %CDT method, a substantial amount of trisialo transferrin was found, which is generally not considered a CDT isoform. Conclusions: The fact that trisialo transferrin is present may generate falsely elevated CDT results and it could at least partly explain the discrepancy between results of the Axis-Shield %CDT assay and HPLC in routine analysis. © 2005 Elsevier B.V. All rights reserved.
  • Bergström, Maria, et al. (författare)
  • Elucidating the selectivity of recombinant forms of Aleuria aurantia lectin using weak affinity chromatography
  • 2012
  • Ingår i: Journal of chromatography. B. - Elsevier. - 1570-0232. ; 885, s. 66-72
  • Tidskriftsartikel (refereegranskat)abstract
    • Aberrant glycosylation is connected to several pathological conditions and lectins are useful tools to characterize glycosylated biomarkers. The Aleuria aurantia lectin (AAL) is of special interest since it interacts with all types of fucosylated saccharides. AAL has been expressed in Escherichia coil as a fully functional recombinant protein. Engineered variants of AAL have been developed with the aim of creating monovalent lectins with more homogenous binding characteristics. Four different forms of AAL were studied in the present work: native AAL purified from A. aurantia mushrooms, recombinant AAL dimer, recombinant AAL monomer and recombinant AAL site 2 (S2-AAL). The affinities of these AAL forms toward a number of saccharides were determined with weak affinity chromatography (WAC). Disaccharides with fucose linked alpha 1-3 to GIcNAc interacted with higher affinity compared to fucose linked alpha 1-6 or alpha 1-4 and the obtained dissociation constants (K-d) were in the range of 10 mu M for all AAL forms. Tetra- and pentasaccharides with fucose in alpha 1-2, alpha 1-3 or alpha 1-4 had K-d values ranging from 0.1 to 7 mM while a large alpha 1-6 fucosylated oligosaccharide had a K-d of about 20 mu M. The recombinant multivalent AAL forms and native AAL exhibited similar affinities toward all saccharides, but S2-AAL had a lower affinity especially regarding a sialic acid containing fucosylated saccharide. It was demonstrated that WAC is a valuable technique in determining the detailed binding profile of the lectins. Specific advantages with WAC include a low consumption of non-labeled saccharides, possibility to analyze mixtures and a simple procedure using standard HPLC equipment.
  • Bergström, Maria, et al. (författare)
  • Lectin Affinity Capillary Electrophoresis in Glycoform Analysis Applying the Partial Filling Technique
  • 2004
  • Ingår i: Journal of Chromatography B. ; 809, s. 323-329
  • Tidskriftsartikel (refereegranskat)abstract
    • The study of protein glycosylation and its significance in biological interactions is a field of growing interest. This work demonstrates a lectin-based separation of protein glycoforms of α1-acid glycoprotein (AGP or orosomucoid) with capillary electrophoresis. Glycoform analysis was performed with a "partial filling technique" with the lectin Concanavalin A (Con A) as affinity ligand. Con A separated human AGP into two peaks, the first peak included AGP glycoforms without biantennary glycans, and the second peak represented the fraction that had one or more biantennary glycans. The applicability of the method was demonstrated with the analysis of AGP from clinical samples and AGP treated with N-glycosidase F. The AGP separation was also used as a reporter system to estimate the dissociation constant (KD) between Con A and a competing sugar. © 2004 Elsevier B.V. All rights reserved.
  • Duong-Thi, Minh-Dao, et al. (författare)
  • Comparison of weak affinity chromatography and surface plasmon resonance in determining affinity of small molecules
  • 2014
  • Ingår i: Analytical Biochemistry. - 0003-2697. ; 461, s. 57-59
  • Tidskriftsartikel (refereegranskat)abstract
    • In this study, we compared affinity data from surface plasmon resonance (SPR) and weak affinity chromatography (WAC), two established techniques for determination of weak affinity (mM-mu M) small molecule-protein interactions. In the current comparison, thrombin was used as target protein. In WAC the affinity constant (K-D) was determined from retention times, and in SPR it was determined by Langmuir isotherm fitting of steady-state responses. Results indicate a strong correlation between the two methods (R-2 = 0.995, P < 0.0001). (C) 2014 Elsevier Inc. All rights reserved.
  • Jungar, C, et al. (författare)
  • Analysis of carbohydrates using liquid chromatography-surface plasmon resonance immunosensing systems
  • 2000
  • Ingår i: Analytical Biochemistry. - 0003-2697. ; 281:2, s. 151-158
  • Tidskriftsartikel (refereegranskat)abstract
    • An immunosensing system based on surface plasmon resonance (SPR) was used for on-line detection and characterization of carbohydrate molecules separated by high-performance liquid chromatography. These analytes, with or without serum, were continuously separated and analyzed in the combined liquid chromatography-surface plasmon resonance (LC-SPR) system. By using weak and readily reversible monoclonal antibodies, the SPR system allowed specific on-line monitoring of the substances. To increase the specificity of the immunosensor, nonrelevant antibodies were used as reference in a serial flow cell. The sensitivity of the LC-SPR system was dependent on molecular weight of the carbohydrate, affinity of binding, and design of the sensor. (C) 2000 Academic Press.
  • Mandenius, Carl-Fredrik, et al. (författare)
  • Monitoring of influenza virus hemagglutinin in process samples using weak affinity ligands and surface plasmon resonance
  • 2008
  • Ingår i: Analytica Chimica Acta. - Elsevier. - 0003-2670. ; 623, s. 66-75
  • Tidskriftsartikel (refereegranskat)abstract
    • Surface plasmon resonance (SPR) was used to screen the interaction between a variety of affinity ligands and hemagglutinin (HA) from human influenza virus, with the aim of identifying low affinity ligands useful for the development of a rapid bioanalytical sensor. Three sialic acid-based structures and four lectins were evaluated as sensor ligands. The sialic acid-based ligands included a natural sialic acid-containing glycoprotein, human α1-acid glycoprotein (α1-AGP), and two synthetic 6′-sialyllactose-conjugates, with varying degree of substitution. The interaction of HA with the four lectin-based ligands, concanavalin A (Con A), wheat germ agglutinin (WGA), Maackia amurensis lectin (MAL), and Sambucus nigra agglutinin (SNA), showed a wide variation of affinity strengths. Affinity and kinetics data were estimated. Strong affinities were observed for Con A, WGA, α1-AGP, and a 6′-sialyllactose-conjugate with a high substitution degree, and low affinities were observed for MAL and a 6′-sialyllactose-conjugate with low substitution.The main objective, to identify a low affinity ligand which could be used for on-line monitoring and product quantification, was met by a 6′-sialyllactose–ovalbumin conjugate that had 0.6 mol ligand per mol carrier protein. The apparent affinity of this ligand was estimated to be 1.5 ± 0.03 μM (KD) on the SPR surface. Vaccine process samples containing HA were analyzed in the range 10–100 μg HA mL−1 and correlated with single-radial immunodiffusion. The coefficient of variation on the same chip was between 0.010 and 0.091.
  • Ohlson, Sten, et al. (författare)
  • Continuous weak-affinity immunosensing
  • 2000
  • Ingår i: Trends in Biotechnology. - 0167-7799. ; 18:2, s. 49-52
  • Tidskriftsartikel (refereegranskat)abstract
    • A multitude of weak biological interactions, either working alone or in concert, occur frequently throughout biological systems. We have used this natural feature of readily reversible interactions as the basis for continuous immunosensing. In a model system, a set of weak monoclonal antibodies directed towards a carbohydrate epitope was studied with the aid of surface plasmon resonance. Because the system requires no regeneration, it can be used as a truly on-line immunosensing device. This principle should have wide application in all areas where there is a need for the continuous evaluation of a molecule.
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