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| 2. |
- Carlsson, Fredrika, et al.
(författare)
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Novel antibody specificities targeting glycoprotein B of cytomegalovirus identified by molecular library technology
- 2009
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Ingår i: New Biotechnology. - : Elsevier BV. - 1876-4347 .- 1871-6784. ; 25:6, s. 429-436
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Tidskriftsartikel (refereegranskat)abstract
- Antibodies provide some protection against cytomegalovirus-mediated disease. All aspects of antibody recognition of important viral antigens are however not fully appreciated. Glycoprotein B (gB), a key protein in the viral membrane, participates in viral infection and it is a component of prototype vaccines. By using combinatorial antibody library and selection technology, novel antibody specificities targeting gB have now been isolated. We define a monoclonal antibody fragment able to recognize site I of antigenic domain (AD) 2, a poorly immunogenic epitope targeted by potent virus-neutralizing antibodies, in a way that is different from the binding of antibodies induced by infection but similar to those induced by vaccination. We also describe the existence of a novel epitope overlapping site I of AD-2 and AD-1, the immunodominant epitope of gB. These specificities, derived by molecular engineering, will be useful for the future assessment of humoral immune responses against this opportunistic viral infection.
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| 4. |
- Gustavsson, Elin, et al.
(författare)
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Surrogate antigens as targets for proteome-wide binder selection
- 2011
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Ingår i: New Biotechnology. - : Elsevier BV. - 1871-6784 .- 1876-4347. ; 28:4, s. 302-311
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Tidskriftsartikel (refereegranskat)abstract
- In the last decade, many initiatives have been taken to develop antibodies for proteome-wide studies, as well as characterization and validation of clinically relevant disease biomarkers. Phage display offers many advantages compared to conventional antibody generation by immunization and hybridoma technology, since it is an unlimited resource of affinity reagents without batch-to-batch variation and is amendable for high throughput. One of the major bottlenecks to proteome-wide binder selection is the limited supply of suitable target antigens representative of the human proteome. Here, we provide proof of principle of using easily accessible, cancer-associated protein epitope signature tags (PrESTs), routinely generated within the Human Protein Atlas project, as surrogate antigens in phage selectionsfor the retrieval of target specific binders. These binders were subsequently tested in western blot, immunohistochemistry and protein microarray application to demonstrate their functionality.
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| 5. |
- Hu, Francis Jingxin, 1986-, et al.
(författare)
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Combination of phage and Gram-positive bacterial display of human antibody repertoires enables isolation of functional high affinity binders
- 2018
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Ingår i: New Biotechnology. - : Elsevier. - 1871-6784 .- 1876-4347. ; 45, s. 80-88
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Tidskriftsartikel (refereegranskat)abstract
- Surface display couples genotype with a surface exposed phenotype and thereby allows screening of gene-encoded protein libraries for desired characteristics. Of the various display systems available, phage display is by far the most popular, mainly thanks to its ability to harbour large size libraries. Here, we describe the first use of a Gram-positive bacterial host for display of a library of human antibody genes which, when combined with phage display, provides ease of use for screening, sorting and ranking by flow cytometry. We demonstrate the utility of this method by identifying low nanomolar affinity scFv fragments towards human epidermal growth factor receptor 2 (HER2). The ranking and performance of the scFv isolated by flow sorting in surface-immobilised form was retained when expressed as soluble scFv and analysed by biolayer interferometry, as well as after expression as full-length antibodies in mammalian cells. We also demonstrate the possibility of using Gram-positive bacterial display to directly improve the affinity of the identified binders via an affinity maturation step using random mutagenesis and flow sorting. This combined approach has the potential for a more complete scan of the antibody repertoire and for affinity maturation of human antibody formats.
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| 6. |
- Johnsson, Ola, et al.
(författare)
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A novel feeding strategy for industrial fed-batch processes based on frequency content analysis
- 2012
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Ingår i: New Biotechnology. - : Elsevier BV. - 1876-4347 .- 1871-6784. ; 29:Supplement, s. 11-11
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Konferensbidrag (refereegranskat)abstract
- Overflow metabolism, i.e. the production of metabolic by-products at a high glycolytic flux, is a recurring problem in fed-batch processes with many types of microorganisms. In the current study, a novel feeding strategy aimed at avoiding process failures due to overflow by-product formation was designed and implemented in a pilot-scale reactor (0.5 m3). The basic principle behind the strategy was to analyze the effects on the dissolved oxygen concentration by periodic variations in the inlet feed rate. The frequency spectrum of the dissolved oxygen signal was used to estimate the proximity of the system to the region where overflow metabolism occurs by examining the content in the relevant frequency range. A control variable based on the measured frequency content was subsequently used to control the feed rate. The only measurement required for this strategy is the dissolved oxygen level in the broth, for which robust, fast and precise probes are widely available in industrial fermentors today. The strategy was successfully implemented in pilot-scale processes for industrial enzyme production using Bacillus licheniformis. It was shown possible to run the process close to the optimal feed rate, indicated by very low amounts of acetate (the overflow metabolite) in the broth. In comparison to a reference strategy the new control strategy resulted in over 10% higher biomass yields.
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| 7. |
- McGinn, Steven, et al.
(författare)
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New Technologies for DNA analysis-A review of the READNA Project.
- 2016
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Ingår i: New Biotechnology. - : Elsevier BV. - 1876-4347 .- 1871-6784.
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Forskningsöversikt (refereegranskat)abstract
- The REvolutionary Approaches and Devices for Nucleic Acid analysis (READNA) project received funding from the European Commission for 4 1/2 years. The objectives of the project revolved around technological developments in nucleic acid analysis. The project partners have discovered, created and developed a huge body of insights into nucleic acid analysis, ranging from improvements and implementation of current technologies to the most promising sequencing technologies that constitute a 3(rd) and 4(th) generation of sequencing methods with nanopores and in situ sequencing, respectively.
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| 10. |
- Säll, Anna, et al.
(författare)
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Advancing the global proteome survey platform by using an oriented single chain antibody fragment immobilization approach
- 2016
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Ingår i: New Biotechnology. - : Elsevier. - 1871-6784 .- 1876-4347. ; 33:5, s. 503-513
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Tidskriftsartikel (refereegranskat)abstract
- Increasing the understanding of a proteome and how its protein composition is affected by for example different diseases, such as cancer, has the potential to improve strategies for early diagnosis and therapeutics. The Global Proteome Survey or GPS is a method that combines mass spectrometry and affinity enrichment with the use of antibodies. The technology enables profiling of complex proteomes in a species independent manner. The sensitivity of GPS, and other methods relying on affinity enrichment, is largely affected by the activity of the exploited affinity reagent. We here present an improvement of the GPS platform by utilizing an antibody immobilization approach which ensures a controlled immobilization process of the antibody to the magnetic bead support. More specifically, we make use of an antibody format that enables site-directed biotinylation and use this in combination with streptavidin coated magnetic beads. The performance of the expanded GPS platform was evaluated by profiling yeast proteome samples. We demonstrate that the oriented antibody immobilization strategy increases the ability of the GPS platform and results in larger fraction of functional antibodies. Additionally, we show that this new antibody format enabled in-solution capture, i.e. immobilization of the antibodies after sample incubation. A workflow has been established that permit the use of an oriented immobilization strategy for the GPS platform.
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