| 1. |
- Brattås, Per Ludvik, et al.
(författare)
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TRIM28 Controls a Gene Regulatory Network Based on Endogenous Retroviruses in Human Neural Progenitor Cells
- 2017
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Ingår i: Cell Reports. - : Elsevier BV. - 2211-1247. ; 18:1, s. 1-11
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Tidskriftsartikel (refereegranskat)abstract
- Endogenous retroviruses (ERVs), which make up 8% of the human genome, have been proposed to participate in the control of gene regulatory networks. In this study, we find a region- and developmental stage-specific expression pattern of ERVs in the developing human brain, which is linked to a transcriptional network based on ERVs. We demonstrate that almost 10,000, primarily primate-specific, ERVs act as docking platforms for the co-repressor protein TRIM28 in human neural progenitor cells, which results in the establishment of local heterochromatin. Thereby, TRIM28 represses ERVs and consequently regulates the expression of neighboring genes. These results uncover a gene regulatory network based on ERVs that participates in control of gene expression of protein-coding transcripts important for brain development.
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| 2. |
- Agaton, Charlotta, et al.
(författare)
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Selective enrichment of monospecific polyclonal antibodies for antibody-based proteomics efforts
- 2004
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Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673 .- 1873-3778. ; 1043, s. 33-40
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Tidskriftsartikel (refereegranskat)abstract
- A high stringency protocol, suitable for systematic purification of polyclonal antibodies, is described. The procedure is designed to allow the generation of target protein-specific antibodies suitable for functional annotation of proteins. Antibodies were generated by immunization with recombinantly produced affinity-tagged target proteins. To obtain stringent recovery of the antibodies, a two-step affinity chromatography principle was devised to first deplete the affinity tag-specific antibodies followed by a second step for affinity capture of the target protein-specific antibodies. An analytical dot-blot array system was developed to analyze the cross-reactivity of the affinity-purified antibodies. The results suggest that the protocol can be used in a highly parallel and automated manner to generate mono-specific polyclonal antibodies for large-scale, antibody-based proteomics efforts, i.e. affinity proteomics.
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| 3. |
- Berglund, Andreas, et al.
(författare)
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15 koncept för bättre ergonomi : Inom äldreomsorg, fysioterapi, däckmontering och varuhantering
- 2015
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Rapport (populärvet., debatt m.m.)abstract
- Den här boken är resultatet av en kurs i ergonomi vid Teknisk design, Luleå tekniska universitet, våren 2015. 15 kursdeltagare har under 10 veckor använt designmetodik och ergonomiska teorier och metoder för att utveckla 15 konceptuella förbättringsförslag baserade på de 4 undersökta kontexterna äldreomsorg, fysioterapi, däckmontering och varuhantering. Fokus för ergonomi inom området teknisk design är att se till att all design, oavsett vilket system det avser, kompletterar människans styrkor och förmågor. Vi ska kort och gott se till att arbetsuppgifter, utrustning, apparater, processer, miljöer och organisationer utformas med människan som utgångspunkt, istället för att tvinga människan att anpassa sig med olika former av överbelastning som möjlig påföljd. För att uppnå detta behöver vi förstå och designa för den variabilitet som är representerad bland oss människor: vi är olika, har olika åldrar, storlek, styrka, kognitiv förmåga, erfarenheter, förväntningar och mål. Att tillämpa ergonomi betyder att studera hur människor interagerar med produkter, processer, miljöer och system för att förbättra dem, dvs. göra dem enklare, säkrare, bekvämare och effektivare att använda. För att kunna göra det behöver vi kunskap om människans förutsättningar och behov. Teknisk design med utgångspunkt och mål i god ergonomi innebär att exempelvis: Att designa produkter och utrustning som är enkla och tillförlitliga att använda med utgångspunkt i kunskap om kognitiv ergonomi, antropometri och belastningsergonomiska och biomekaniska analyserAtt designa säkra och effektiva tillverkningsprocesser med utgångspunkt i kunskap om kognitiv ergonomi och belastningsergonomiska analyserAtt designa organisationer utifrån kunskap om arbetslivsfysiologi och organisationsergonomiAtt designa arbetsuppgifter utifrån kunskap om kognitiv ergonomi, biomekanik och belastningsergonomiska analyserAtt designa enkla och användarvänliga gränssnitt med utgångspunkt i kognitiv ergonomiErgonomisk anpassning av en produkt eller en arbetsmiljö kan exempelvis handla om att se till att människan inte använder kroppen felaktigt. Det kan handla om fysisk belastning när en uppgift utförs, såväl som sensorisk input från olika system eller psykosocial belastning i form av stress. Det handlar om att utveckla kunskaper om människans begränsningar och förmågor, vilket ger bättre förutsättningar att bidra till användarvänliga lösningar. Det i sin tur bidrar till säkerhet och användarvänlighet och i slutändan att alla produkter, system och miljöer i vår omvärld fungerar väl för människan – det är hållbar utveckling om något. I kursen Ergonomi 2 vid civilingenjörsutbildningen Teknisk design, Luleå tekniska universitet, ingår en projektuppgift. Den syftar till att få fördjupad förståelse inom ergonomi genom att tillämpa kunskap och metoder i ett designprojekt för en verklig situation. Våren 2015 omfattade projektuppgiften att enanalys av valfri kontext, med syfte att förstå problem och utmaningar i den miljö, det sammanhang, den situation och för de personer som var berörda. Inledningsvis arbetade kursdeltagarna i grupper bestående av 3-4 personer, för att sedan gå in i en konceptutvecklingsfas individuellt. Det innebar att kursdeltagarna kunde genomföra ergonomiska analyser gemensamt och sedan utveckla konceptuella lösningar på egen hand. Det resulterade i att kursdeltagarna utvecklade tämligen olika lösningar, även om de haft en gemensam utgångspunkt. Bokens kapitel omfattar en beskrivning av respektive kontext följt av de konceptförslag som kursdeltagarna utvecklade. Som lärare är det alltid extra roligt när kursdeltagare är motiverade och engagerade inför projektuppgifter. Vår förhoppning är att det engagemanget ska framgå på följande sidor och att koncepten ska ge inspiration till att förbättra ergonomin i våra vardagsliv. Åsa Wikberg Nilsson, Therese Öhrling, Lars Sundström, Agneta Larsson och Ulrik RöijezonTeknisk design Luleå tekniska universitet, Augusti 2015
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| 4. |
- Bonvicini, Gillian, et al.
(författare)
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Comparing in vitro affinity measurements of antibodies to TfR1 : Surface plasmon resonance versus on-cell affinity
- 2024
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Ingår i: Analytical Biochemistry. - : Elsevier. - 0003-2697 .- 1096-0309. ; 686
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Tidskriftsartikel (refereegranskat)abstract
- Despite years of utilizing the transferrin receptor 1 (TfR1) to transport large biomolecules into the brain, there is no consensus on how to optimally measure affinity to it. The aim of this study was to compare different methods for measuring the affinities of anti-TfR1 antibodies.Antibodies 15G11, OX26 and 8D3 are known to successfully carry large biologics across the blood-brain barrier in humans, rats, and mice, respectively. The affinity to their respective species of TfR1 was measured with different surface plasmon resonance setups in Biacore and an on-cell assay.When the antibody was captured and TfR1 was the analyte, the dissociation in Biacore was very slow. The dissociation was faster when the antibody was the analyte and TfR1 was the ligand. The Biacore setup with capture of N-terminal FLAG-tag TfR1 yielded the most similar apparent affinities as the cell assay.In conclusion, it is important to evaluate assay parameters including assay orientation, surface capture method, and antibody format when comparing binding kinetics for TfR1 antibodies. Although it seems possible to determine relative affinities of TfR1 antibodies using the methods described here, both the FLAG-tag TfR1 capture setup and cell assays likely yield apparent affinities that are most translatable in vivo.
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| 5. |
- Falk, Ronny, et al.
(författare)
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A high-stringency proteomics concept aimed for generation of antibodies specific for cDNAencoded proteins
- 2002
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Ingår i: Biotechnology and applied biochemistry. - 0885-4513 .- 1470-8744. ; 35:2, s. 75-82
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Tidskriftsartikel (refereegranskat)abstract
- A novel dual bacterial expression system, designed for high-throughput generation of antibodies specific for cDNA-encoded proteins, is presented. The concept involves parallel expression of cDNA-encoded proteins, in two vector systems, as fusions with two different tags, both enabling single-step affinity purification under denaturing conditions. One of the fusion tags includes a portion with documented immunopotentiating effect to stimulate antibody production, and the generated fusion proteins are used to elicit antibodies. The second fusion protein is used in an immobilized form as an affinity ligand to enrich, from the generated antisera, antibodies with selective reactivity to the cDNA-encoded part. To evaluate the system, five cDNA clones from a mouse testis cDNA library were expressed, and antibodies to these cDNA-encoded proteins were generated, enriched and used in blotting procedures to determine expression patterns for the native proteins corresponding to the cDNAs. The five antibody preparations showed specific reactivity to the corresponding recombinant cDNA-encoded proteins, and three of the five antibodies gave specific staining in Western-blot screening of various cell types and tissue homogenates. When the same five cDNAs were processed and analysed using a single-vector method, antibodies with a more non-specific staining were generated. We thus conclude that the presented dual-vector method offers a highly stringent strategy for generation of monospecific polyclonal antibodies.
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| 6. |
- Falk, Ronny, et al.
(författare)
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An improved dual-expression concept, generating high-quality antibodies for proteomics research
- 2003
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Ingår i: Biotechnology and applied biochemistry. - : Wiley. - 0885-4513 .- 1470-8744. ; 38, s. 231-239
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Tidskriftsartikel (refereegranskat)abstract
- A novel, improved dual bacterial-expression system, designed for large-scale generation of high-quality polyclonal antibody preparations intended for proteomics research, is presented. The concept involves parallel expression of cDNA-encoded proteins, as a fusion with two different tags in two separate vector systems. Both systems enable convenient blotting procedures for expression screening on crude bacterial cell cultures and single-step affinity purification under denaturing conditions. One of the fusion proteins is used to elicit antibodies, and the second fusion protein is used in an immobilized form as an affinity ligand to enrich antibodies with selective reactivity to the cDNA-encoded part, common for the two fusion proteins. To evaluate the system, four cDNA clones from putative nuclear proteins from the non-biting midge Chironomus tentans were expressed. Antibodies to these cDNA-encoded proteins were generated, enriched and used in blotting and immunofluorescence procedures to determine expression patterns for the native proteins corresponding to the cDNAs. The four antibody preparations showed specific reactivity to the corresponding recombinant cDNA-encoded proteins, and three of the four antibodies gave specific staining in Western-blot analysis of nuclear cell extracts. Furthermore, two of the antibody preparations gave specific staining in immunofluorescence analysis of C. tentans cells. We conclude that the dual-vector concept presented offers a highly stringent strategy for the generation of monospecific polyclonal antibodies, which are useful in proteomics research.
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| 7. |
- Falk, Ronny, et al.
(författare)
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Approaches for systematic proteome exploration
- 2007
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Ingår i: Biomolecular Engineering. - : Elsevier BV. - 1389-0344 .- 1878-559X. ; 24:2, s. 155-168
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Forskningsöversikt (refereegranskat)abstract
- With the completion of the human genome project (HUGO) during recent years, gene function, protein abundance and expression patterns in tissues and cell types have emerged as central areas for the scientific community. A mapped human proteome will extend the value of the genome sequence and large-scale efforts aiming at elucidating protein localization, abundance and function are invaluable for biomarker and drug discovery. This research area, termed proteomics, is more demanding than any genome sequencing effort and to perform this on a wide scale is a highly diverse task. Therefore, the proteornics field employs a range of methods to examine different aspects of proteomics including protein localization, protein-protein interactions, posttranslational modifications and alteration of protein composition (e.g. differential expression) in tissues and body fluids. Here, some of the most commonly used methods, including chromatographic separations together with mass spectrometry and a number of affinity proteomics concepts are discussed and exemplified.
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| 8. |
- Falk, Ronny, et al.
(författare)
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Intervju med journalist Bengt Larsson från Kiruna
- 1981
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Annan publikation (populärvet., debatt m.m.)abstract
- Interview with Bengt Larsson from Kiruna. The interview was conducted by Ronny Falk, Pia Gillberg and Inger Hjelm, students of the Swedish School of Library and Information Science in Borås, in 1981.
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| 9. |
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| 10. |
- Falk, Ronny, 1970-
(författare)
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Systems enabling antibody-mediated proteomics research
- 2006
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- As many genome sequencing efforts today are completed, we are now provided with the genetic maps for several organisms, including man. With these maps at hand, the scientific focus is now shifting towards investigations of the functionality of proteins. This task is even more challenging than the genomic field since proteins, in contrast to DNA, do not allow themselves to be specifically probed or amplified by easy and generic methods. However, to achieve knowledge regarding protein function, useful information includes where, when and how much certain proteins are expressed in an organism. Such information can be obtained if protein-specific binding molecules are available as tools. One such class of target specific binders are the antibody molecules, traditionally employed in a broad variety of biotechnical applications, including protein localization studies on both cellular and sub cellular levels.In a first serie of studies, new methodology for recombinant production and purification of antigens for generation of antibodies via immunization routes were investigated. Parallel affinity gene fusion-based expression systems were used for evaluation of different concepts for production of antigen and post-immunization antibody purification. Carefully designed protein antigens from different organisms were produced and used to raise antisera which were affinity purified on their respective antigens to obtain highly specific polyclonal antibodies (monospecific antibodies). One of the constructed expression systems includes an affinity handle, ZSPA-1, previously selected from a combinatorial protein library for its capacity to selectively bind protein A. This allows for convenient, non IgG-dependent, affinity purification of proteins on conventional protein A resins.A strategy where highly target specific antibody preparations could be affinity purified in a more streamlined setup is also presented. By this strategy it was possible to fractionate antibodies showing reactivity to different parts of the antigen into separate fractions. This resulted in affinity purified antibodies showing monospecific but still multi-epitope reactivity. Purified monospecific antibodies were used in different studies including Western blot immunofluorescence and recovery applications. For affinity purification of endogenous target from its native surrounding a selective elution strategy where the recombinant antigen was used to competitively elute the captured target was developed.
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