| 1. |
- Nilsson, Christian, et al.
(författare)
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Cationic and anionic lipid-based nanoparticles in CEC for protein separation.
- 2010
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Ingår i: Electrophoresis. - : Wiley. - 0173-0835. ; 31:11, s. 1773-1779
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Tidskriftsartikel (refereegranskat)abstract
- The development of new separation techniques is an important task in protein science. Herein, we describe how anionic and cationic lipid-based liquid crystalline nanoparticles can be used for protein separation. The potential of the suggested separation methods is demonstrated on green fluorescent protein (GFP) samples for future use on more complex samples. Three different CEC-LIF approaches for protein separation are described. (i) GFP and GFP N212Y, which are equally charged, were separated with high resolution by using anionic nanoparticles suspended in the electrolyte and adsorbed to the capillary wall. (ii) High efficiency (800 000 plates/m) and peak capacity were demonstrated separating GFP samples from Escherichia coli with cationic nanoparticles suspended in the electrolyte and adsorbed to the capillary wall. (iii) Three single amino-acid-substituted GFP variants were separated with high resolution using an approach based on a physical attached double-layer coating of cationic and anionic nanoparticles combined with anionic lipid nanoparticles suspended in the electrolyte. The soft and porous lipid-based nanoparticles were synthesized by a one-step procedure based on the self-assembly of lipids, and were biocompatible with a large surface-to-volume ratio. The methodology is still under development and the optimization of the nanoparticle chemistry and separation conditions can further improve the separation system. In contrast to conventional LC, a new interaction phase is introduced for every analysis, which minimizes carry-over and time-consuming column regeneration.
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| 2. |
- Nilsson, Christian, et al.
(författare)
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Hydrophobic Interaction Capillary Electrochromatography of Protein Mutants. Use of Lipid-Based Liquid Crystalline Nanoparticles as Pseudostationary Phase.
- 2009
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Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 1520-6882 .- 0003-2700. ; 81:1, s. 315-321
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Tidskriftsartikel (refereegranskat)abstract
- Nanoparticle-based hydrophobic interaction-capillary electrochromatography was utilized for separation of proteins with similar mass-to-charge ratio at neutral pH without organic modifier. Lipid-based liquid crystalline nanoparticles were prepared and used as pseudostationary phase, benefiting from their high biocompatibility, ease of preparation, and suspension stability at high concentrations. Use of laser-induced fluorescence enabled detection at high nanoparticle concentrations. Green fluorescent protein (GFP) and mutants of GFP harboring single or double amino acid substitutions with the same charge were separated in the described system but not in conventional capillary electrophoresis. Separation was achieved by increasing the salt concentration to promote hydrophobic interactions by shielding of the repulsive electrostatic interactions. In addition, the method was adapted to a capillary with an effective length of 6.7 cm, enabling fast separations and future applications on chip.
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| 3. |
- Nilsson, Christian, et al.
(författare)
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Nanoparticle-based capillary electroseparation of proteins in polymer capillaries under physiological conditions.
- 2010
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Ingår i: Electrophoresis. - : Wiley. - 0173-0835 .- 1522-2683. ; 31:3, s. 459-464
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Tidskriftsartikel (refereegranskat)abstract
- Totally porous lipid-based liquid crystalline nanoparticles were used as pseudostationary phase for capillary electroseparation with LIF detection of proteins at physiological conditions using unmodified cyclic olefin copolymer capillaries (Topas, 6.7 cm effective length). In the absence of nanoparticles, i.e. in CE mode, the protein samples adsorbed completely to the capillary walls and could not be recovered. In contrast, nanoparticle-based capillary electroseparation resolved green fluorescent protein from several of its impurities within 1 min. Furthermore, a mixture of native green fluorescent protein and two of its single-amino-acid-substituted variants was separated within 2.5 min with efficiencies of 400 000 plates/m. The nanoparticles prevent adsorption by introducing a large interacting surface and by obstructing the attachment of the protein to the capillary wall. A one-step procedure based on self-assembly of lipids was used to prepare the nanoparticles, which benefit from their biocompatibility and suspension stability at high concentrations. An aqueous tricine buffer at pH 7.5 containing lipid-based nanoparticles (2% w/w) was used as electrolyte, enabling separation at protein friendly conditions. The developed capillary-based method facilitates future electrochromatography of proteins on polymer-based microchips under physiological conditions and enables the initial optimization of separation conditions in parallel to the chip development.
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| 4. |
- Nilsson, Christian, et al.
(författare)
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Nanoparticle-based pseudostationary phases in CEC: A breakthrough in protein analysis?
- 2011
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Ingår i: Electrophoresis. - : Wiley. - 0173-0835. ; 32:10, s. 1141-1147
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Forskningsöversikt (refereegranskat)abstract
- This review focuses on major trends in nanoparticle-based pseudostationary phase (PSP) CEC since the publication of our previous reviews within nanoparticle-based CEC [Nilsson, C., et al., Electrophoresis 2006, 27, 76-83; Nilsson, C., et al., J. Chromatogr. A 2007, 1168, 212-224.]. Special attention is given to the development toward protein analysis, which is driven by the strong emergence of protein drug development in the pharmaceutical industry. Furthermore, we discuss the development in coupling different detection techniques with nanoparticle-based PSP CEC, which were originally predicted to be particularly cumbersome. However, at present, direct UV, LIF and ESI-MS have been used without any severe complications. Different types of nanoparticles used as PSP during the period include gold nanoparticles, carbon nanostructures and lipid-based nanoparticles. New materials (for example, different types of carbon nanostructures and self assembled lipid-based nanostructures) are a strong driving force for development in separation science. Finally, future trends in nanoparticles-based CEC are envisioned.
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| 5. |
- Nilsson, Christian, et al.
(författare)
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Use of nanoparticles in capillary and microchip electrochromatography
- 2007
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Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 1168:1-2, s. 212-224
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Forskningsöversikt (refereegranskat)abstract
- Applications of nanoparticles are of rising interest in separation science, due to their favorable surface-to-volume ratio as well as their applicability in miniaturization. A stationary phase with large surface area in combination with an electroosmotic flow-driven system has great potential in a highly efficient separation system. This review covers the use of various nanoparticles as stationary or pseudostationary phase in capillary and microchip electrochromatography. The use of nanoparticles in pseudostationary phase capillary electrochromatography and open-tubular capillary electrochromatography are thoroughly discussed. The stationary and pseudostationary phases that are described include polymer nanoparticles, gold nanoparticles, silica nanoparticles, fullerenes and carbon nanotubes. (c) 2007 Elsevier B.V. All rights reserved.
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| 6. |
- Tomic, Tajana Tesan, et al.
(författare)
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MYO5B mutations in pheochromocytoma/paraganglioma promote cancer progression
- 2020
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Ingår i: PLOS Genetics. - : Public Library of Science. - 1553-7390 .- 1553-7404. ; 16:6
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Tidskriftsartikel (refereegranskat)abstract
- Identification of additional cancer-associated genes and secondary mutations driving the metastatic progression in pheochromocytoma and paraganglioma (PPGL) is important for subtyping, and may provide optimization of therapeutic regimens. We recently reported novel recurrent nonsynonymous mutations in the MYO5B gene in metastatic PPGL. Here, we explored the functional impact of these MYO5B mutations, and analyzed MYO5B expression in primary PPGL tumor cases in relation to mutation status. Immunohistochemistry and mRNA expression analysis in 30 PPGL tumors revealed an increased MYO5B expression in metastatic compared to non-metastatic cases. In addition, subcellular localization of MYO5B protein was altered from cytoplasmic to membranous in some metastatic tumors, and the strongest and most abnormal expression pattern was observed in a paraganglioma harboring a somatic MYO5B:p.G1611S mutation. In addition to five previously discovered MYO5B mutations, the present study of 30 PPGL (8 previous and 22 new samples) also revealed two, and hence recurrent, mutations in the gene paralog MYO5A. The three MYO5B missense mutations with the highest prediction scores (p.L587P, p.G1611S and p.R1641C) were selected and functionally validated using site directed mutagenesis and stable transfection into human neuroblastoma cells (SK-N-AS) and embryonic kidney cells (HEK293). In vitro analysis showed a significant increased proliferation rate in all three MYO5B mutated clones. The two somatically derived mutations, p.L587P and p.G1611S, were also found to increase the migration rate. Expression analysis of MYO5B mutants compared to wild type clones, demonstrated a significant enrichment of genes involved in migration, proliferation, cell adhesion, glucose metabolism, and cellular homeostasis. Our study validates the functional role of novel MYO5B mutations in proliferation and migration, and suggest the MYO5-pathway to be involved in the malignant progression in some PPGL tumors. © 2020 Tomic et al.
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| 7. |
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| 8. |
- Moller, Anders Pape, et al.
(författare)
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Effects of interspecific coexistence on laying date and clutch size in two closely related species of hole-nesting birds
- 2018
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Ingår i: Journal of Animal Ecology. - : WILEY. - 0021-8790 .- 1365-2656. ; 87:6, s. 1738-1748
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Tidskriftsartikel (refereegranskat)abstract
- Coexistence between great tits Parus major and blue tits Cyanistes caeruleus, but also other hole-nesting taxa, constitutes a classic example of species co-occurrence resulting in potential interference and exploitation competition for food and for breeding and roosting sites. However, the spatial and temporal variations in coexistence and its consequences for competition remain poorly understood. We used an extensive database on reproduction in nest boxes by great and blue tits based on 87 study plots across Europe and Northern Africa during 1957-2012 for a total of 19,075 great tit and 16,729 blue tit clutches to assess correlative evidence for a relationship between laying date and clutch size, respectively, and density consistent with effects of intraspecific and interspecific competition. In an initial set of analyses, we statistically controlled for a suite of site-specific variables. We found evidence for an effect of intraspecific competition on blue tit laying date (later laying at higher density) and clutch size (smaller clutch size at higher density), but no evidence of significant effects of intraspecific competition in great tits, nor effects of interspecific competition for either species. To further control for site-specific variation caused by a range of potentially confounding variables, we compared means and variances in laying date and clutch size of great and blue tits among three categories of difference in density between the two species. We exploited the fact that means and variances are generally positively correlated. If interspecific competition occurs, we predicted a reduction in mean and an increase in variance in clutch size in great tit and blue tit when density of heterospecifics is higher than the density of conspecifics, and for intraspecific competition, this reduction would occur when density of conspecifics is higher than the density of heterospecifics. Such comparisons of temporal patterns of means and variances revealed evidence, for both species, consistent with intraspecific competition and to a smaller extent with interspecific competition. These findings suggest that competition associated with reproductive behaviour between blue and great tits is widespread, but also varies across large spatial and temporal scales.
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| 9. |
- Pape Møller, Anders, et al.
(författare)
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Interaction of climate change with effects of conspecific and heterospecific density on reproduction
- 2020
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Ingår i: Oikos. - : Wiley. - 0030-1299 .- 1600-0706. ; 129:12, s. 1807-1819
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Tidskriftsartikel (refereegranskat)abstract
- We studied the relationship between temperature and the coexistence of great tit Parus major and blue tit Cyanistes caeruleus, breeding in 75 study plots across Europe and North Africa. We expected an advance in laying date and a reduction in clutch size during warmer springs as a general response to climate warming and a delay in laying date and a reduction in clutch size during warmer winters due to density‐dependent effects. As expected, as spring temperature increases laying date advances and as winter temperature increases clutch size is reduced in both species. Density of great tit affected the relationship between winter temperature and laying date in great and blue tit. Specifically, as density of great tit increased and temperature in winter increased both species started to reproduce later. Density of blue tit affected the relationship between spring temperature and blue and great tit laying date. Thus, both species start to reproduce earlier with increasing spring temperature as density of blue tit increases, which was not an expected outcome, since we expected that increasing spring temperature should advance laying date, while increasing density should delay it cancelling each other out. Climate warming and its interaction with density affects clutch size of great tits but not of blue tits. As predicted, great tit clutch size is reduced more with density of blue tits as temperature in winter increases. The relationship between spring temperature and density on clutch size of great tits depends on whether the increase is in density of great tit or blue tit. Therefore, an increase in temperature negatively affected the coexistence of blue and great tits differently in both species. Thus, blue tit clutch size was unaffected by the interaction effect of density with temperature, while great tit clutch size was affected in multiple ways by these interactions terms.
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| 10. |
- Moller, Anders Pape, et al.
(författare)
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Clutch-size variation in Western Palaearctic secondary hole-nesting passerine birds in relation to nest box design
- 2014
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Ingår i: Methods in Ecology and Evolution. - 2041-210X. ; 5:4, s. 353-362
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Tidskriftsartikel (refereegranskat)abstract
- Secondary hole-nesting birds that do not construct nest holes themselves and hence regularly breed in nest boxes constitute important model systems for field studies in many biological disciplines with hundreds of scientists and amateurs involved. Those research groups are spread over wide geographic areas that experience considerable variation in environmental conditions, and researchers provide nest boxes of varying designs that may inadvertently introduce spatial and temporal variation in reproductive parameters. We quantified the relationship between mean clutch size and nest box size and material after controlling for a range of environmental variables in four of the most widely used model species in the Western Palaearctic: great tit Parus major, blue tit Cyanistes caeruleus, pied flycatcher Ficedula hypoleuca and collared flycatcher F.albicollis from 365 populations and 79610 clutches. Nest floor area and nest box material varied non-randomly across latitudes and longitudes, showing that scientists did not adopt a random box design. Clutch size increased with nest floor area in great tits, but not in blue tits and flycatchers. Clutch size of blue tits was larger in wooden than in concrete nest boxes. These findings demonstrate that the size of nest boxes and material used to construct nest boxes can differentially affect clutch size in different species. The findings also suggest that the nest box design may affect not only focal species, but also indirectly other species through the effects of nest box design on productivity and therefore potentially population density and hence interspecific competition.
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