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  • 301.
    Mardinoglu, Adil
    et al.
    KTH, Centra, Science for Life Laboratory, SciLifeLab. Chalmers Univ Technol, Dept Biol & Biol Engn, Gothenburg, Sweden..
    Boren, Jan
    Univ Gothenburg, Dept Mol & Clin Med, Gothenburg, Sweden.;Sahlgrens Univ Hosp, Gothenburg, Sweden..
    Smith, Ulf
    Univ Gothenburg, Dept Mol & Clin Med, Gothenburg, Sweden.;Sahlgrens Univ Hosp, Gothenburg, Sweden..
    Uhlén, Mathias
    KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Nielsen, Jens
    KTH, Centra, Science for Life Laboratory, SciLifeLab. Chalmers Univ Technol, Dept Biol & Biol Engn, Gothenburg, Sweden..
    Systems biology in hepatology: approaches and applications2018Inngår i: Nature Reviews. Gastroenterology & Hepatology, ISSN 1759-5045, E-ISSN 1759-5053, Vol. 15, nr 6, s. 365-377Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Detailed insights into the biological functions of the liver and an understanding of its crosstalk with other human tissues and the gut microbiota can be used to develop novel strategies for the prevention and treatment of liver-associated diseases, including fatty liver disease, cirrhosis, hepatocellular carcinoma and type 2 diabetes mellitus. Biological network models, including metabolic, transcriptional regulatory, protein-protein interaction, signalling and co-expression networks, can provide a scaffold for studying the biological pathways operating in the liver in connection with disease development in a systematic manner. Here, we review studies in which biological network models were used to integrate multiomics data to advance our understanding of the pathophysiological responses of complex liver diseases. We also discuss how this mechanistic approach can contribute to the discovery of potential biomarkers and novel drug targets, which might lead to the design of targeted and improved treatment strategies. Finally, we present a roadmap for the successful integration of models of the liver and other human tissues with the gut microbiota to simulate whole-body metabolic functions in health and disease.

  • 302. Mardinoglu, Adil
    et al.
    Kampf, Caroline
    Asplund, Anna
    Fagerberg, Linn
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Hallström, Björn M.
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Edlund, Karolina
    Blüher, Matthias
    Pontén, Fredrik
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Nielsen, Jens
    KTH, Skolan för bioteknologi (BIO), Genteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab. Chalmers University of Technology, Sweden.
    Defining the Human Adipose Tissue Proteome To Reveal Metabolic Alterations in Obesity2014Inngår i: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 13, nr 11, s. 5106-5119Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    White adipose tissue (WAT) has a major role in the progression of obesity. Here, we combined data from RNA-Seq and antibody-based immunohistochemistry to describe the normal physiology of human WAT obtained from three female subjects and explored WAT-specific genes by comparing WAT to 26 other major human tissues. Using the protein evidence in WAT, we validated the content of a genome-scale metabolic model for adipocytes. We employed this high-quality model for the analysis of subcutaneous adipose tissue (SAT) gene expression data obtained from subjects included in the Swedish Obese Subjects Sib Pair study to reveal molecular differences between lean and obese individuals. We integrated SAT gene expression and plasma metabolomics data, investigated the contribution of the metabolic differences in the mitochondria of SAT to the occurrence of obesity, and eventually identified cytosolic branched-chain amino acid (BCAA) transaminase 1 as a potential target that can be used for drug development. We observed decreased glutaminolysis and alterations in the BCAAs metabolism in SAT of obese subjects compared to lean subjects. We also provided mechanistic explanations for the changes in the plasma level of BCAAs, glutamate, pyruvate, and alpha-ketoglutarate in obese subjects. Finally, we validated a subset of our model-based predictions in 20 SAT samples obtained from 10 lean and 10 obese male and female subjects.

  • 303. Mardinoglu, Adil
    et al.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Nielsen, Jens
    The use of Genome-scale metabolic models for drug target and biomarker identification2014Inngår i: New Biotechnology, ISSN 1871-6784, E-ISSN 1876-4347, Vol. 31, s. S49-S49Artikkel i tidsskrift (Annet vitenskapelig)
  • 304.
    Mardinoglu, Adil
    et al.
    KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Ural, Dilek
    Koc Univ, Sch Med, TR-34450 Istanbul, Turkey..
    Zeybel, Mujdat
    Koc Univ, Sch Med, Dept Gastroenterol & Hepatol, TR-34450 Istanbul, Turkey..
    Yuksel, Hatice Hilal
    KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Uhlén, Mathias
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för bioteknologi (BIO), Centra, Albanova VinnExcellence Center for Protein Technology, ProNova. KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap.
    Boren, Jan
    Univ Gothenburg, Dept Mol & Clin Med, S-41345 Gothenburg, Sweden.;Sahlgrenska Univ Hosp Gothenburg, S-41345 Gothenburg, Sweden..
    The Potential Use of Metabolic Cofactors in Treatment of NAFLD2019Inngår i: Nutrients, ISSN 2072-6643, E-ISSN 2072-6643, Vol. 11, nr 7, artikkel-id 1578Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Non-alcoholic fatty liver disease (NAFLD) is caused by the imbalance between lipid deposition and lipid removal from the liver, and its global prevalence continues to increase dramatically. NAFLD encompasses a spectrum of pathological conditions including simple steatosis and non-alcoholic steatohepatitis (NASH), which can progress to cirrhosis and liver cancer. Even though there is a multi-disciplinary effort for development of a treatment strategy for NAFLD, there is not an approved effective medication available. Single or combined metabolic cofactors can be supplemented to boost the metabolic processes altered in NAFLD. Here, we review the dosage and usage of metabolic cofactors including l-carnitine, Nicotinamide riboside (NR), l-serine, and N-acetyl-l-cysteine (NAC) in human clinical studies to improve the altered biological functions associated with different human diseases. We also discuss the potential use of these substances in treatment of NAFLD and other metabolic diseases including neurodegenerative and cardiovascular diseases of which pathogenesis is linked to mitochondrial dysfunction.

  • 305. Mardinoglu, Adil
    et al.
    Ågren, Rasmus
    Kampf, Caroline
    Asplund, Anna
    Nookaew, Intawat
    Jacobson, Peter
    Walley, Andrew J.
    Froguel, Philippe
    Carlsson, Lena M.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Nielsen, Jens
    Integration of clinical data with a genome-scale metabolic model of the human adipocyte2013Inngår i: Molecular Systems Biology, ISSN 1744-4292, E-ISSN 1744-4292, Vol. 9, s. 649-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    We evaluated the presence/absence of proteins encoded by 14 077 genes in adipocytes obtained from different tissue samples using immunohistochemistry. By combining this with previously published adipocyte-specific proteome data, we identified proteins associated with 7340 genes in human adipocytes. This information was used to reconstruct a comprehensive and functional genome-scale metabolic model of adipocyte metabolism. The resulting metabolic model, iAdipocytes1809, enables mechanistic insights into adipocyte metabolism on a genome-wide level, and can serve as a scaffold for integration of omics data to understand the genotype-phenotype relationship in obese subjects. By integrating human transcriptome and fluxome data, we found an increase in the metabolic activity around androsterone, ganglioside GM2 and degradation products of heparan sulfate and keratan sulfate, and a decrease in mitochondrial metabolic activities in obese subjects compared with lean subjects. Our study hereby shows a path to identify new therapeutic targets for treating obesity through combination of high throughput patient data and metabolic modeling.

  • 306. Mardinoglu, Adil
    et al.
    Ågren, Rasmus
    Kampf, Caroline
    Asplund, Anna
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Nielsen, Jens
    KTH, Skolan för bioteknologi (BIO), Genteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Genome-scale metabolic modelling of hepatocytes reveals serine deficiency in patients with non-alcoholic fatty liver disease2014Inngår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 5, s. 3083-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Several liver disorders result from perturbations in the metabolism of hepatocytes, and their underlying mechanisms can be outlined through the use of genome-scale metabolic models (GEMs). Here we reconstruct a consensus GEM for hepatocytes, which we call iHepatocytes2322, that extends previous models by including an extensive description of lipid metabolism. We build iHepatocytes2322 using Human Metabolic Reaction 2.0 database and proteomics data in Human Protein Atlas, which experimentally validates the incorporated reactions. The reconstruction process enables improved annotation of the proteomics data using the network centric view of iHepatocytes2322. We then use iHepatocytes2322 to analyse transcriptomics data obtained from patients with non-alcoholic fatty liver disease. We show that blood concentrations of chondroitin and heparan sulphates are suitable for diagnosing non-alcoholic steatohepatitis and for the staging of non-alcoholic fatty liver disease. Furthermore, we observe serine deficiency in patients with NASH and identify PSPH, SHMT1 and BCAT1 as potential therapeutic targets for the treatment of non-alcoholic steatohepatitis.

  • 307. Mathivanan, Suresh
    et al.
    Ahmed, Mukhtar
    Ahn, Natalie G.
    Hainard, Alexandre
    Trinidad, Jonathan C.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Pandey, Akhilesh
    Human Proteinpedia enables sharing of human protein data2008Inngår i: Nature Biotechnology, ISSN 1087-0156, E-ISSN 1546-1696, Vol. 26, nr 2, s. 164-167Artikkel i tidsskrift (Fagfellevurdert)
  • 308. Matic, L. P.
    et al.
    Iglesias, Maria Jesus
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Vesterlund, M.
    Lengquist, M.
    Hong, Mun-Gwan
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Saieed, S.
    Sanchez-Rivera, Laura
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Berg, M.
    Razuvaev, A.
    Kronqvist, M.
    Lund, K.
    Caidahl, K.
    Gillgren, P.
    Pontén, F.
    Uhlén, Mathias
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Schwenk, Jochen M.
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Hansson, G. K.
    Paulsson-Berne, G.
    Fagman, E.
    Roy, J.
    Hultgren, R.
    Bergström, G.
    Lehtiö, J.
    Odeberg, Jacob
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Hedin, U.
    Novel Multiomics Profiling of Human Carotid Atherosclerotic Plaques and Plasma Reveals Biliverdin Reductase B as a Marker of Intraplaque Hemorrhage2018Inngår i: JACC: Basic to Translational Science, ISSN 2452-302X, Vol. 3, nr 4, s. 464-480Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Clinical tools to identify individuals with unstable atherosclerotic lesions are required to improve prevention of myocardial infarction and ischemic stroke. Here, a systems-based analysis of atherosclerotic plaques and plasma from patients undergoing carotid endarterectomy for stroke prevention was used to identify molecular signatures with a causal relationship to disease. Local plasma collected in the lesion proximity following clamping prior to arteriotomy was profiled together with matched peripheral plasma. This translational workflow identified biliverdin reductase B as a novel marker of intraplaque hemorrhage and unstable carotid atherosclerosis, which should be investigated as a potential predictive biomarker for cardiovascular events in larger cohorts.

  • 309. Mattsson, Johanna S. M.
    et al.
    Svensson, Maria A.
    Hallström, Björn
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Koyi, Hirsh
    Branden, Eva
    Brunnstrom, Hans
    Edlund, Karolina
    Ekman, Simon
    La Fleur, Linnea
    Grinberg, Marianna
    Rahnenfuehrer, Joerg
    Jirstrom, Karin
    Ponten, Fredrik
    Karlsson, Mats G.
    Karlsson, Christina
    Helenius, Gisela
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Botling, Johan
    Micke, Patrick
    ALK Rearrangements in Non-Small Cell Lung Cancer: Comprehensive Integration of Genomic, Gene Expression and Protein Analysis2015Inngår i: Journal of Thoracic Oncology, ISSN 1556-0864, E-ISSN 1556-1380, Vol. 10, nr 9, s. S298-S298Artikkel i tidsskrift (Annet vitenskapelig)
  • 310.
    Mikus, Maria
    et al.
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Drobin, Kimi
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Gry, Marcus
    KTH.
    Bachmann, J.
    Lindberg, J.
    Yimer, G.
    Aklillu, E.
    Makonnen, E.
    Aderaye, G.
    Roach, J.
    Fier, I.
    Kampf, C.
    Göpfert, J.
    Perazzo, H.
    Poynard, T.
    Stephens, C.
    Andrade, R. J.
    Lucena, M. I.
    Arber, N.
    Uhlén, Mattias
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Watkins, P. B.
    Schwenk, Jochen M
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Nilsson, P. Anders
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Schuppe-Koistinen, I.
    Elevated levels of circulating CDH5 and FABP1 in association with human drug-induced liver injury2016Inngår i: Liver international (Print), ISSN 1478-3223, E-ISSN 1478-3231, Vol. 37, nr 1, s. 132-140Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background & Aims: The occurrence of drug-induced liver injury (DILI) is a major issue in all phases of drug development. To identify novel biomarker candidates associated with DILI, we utilised an affinity proteomics strategy, where antibody suspension bead arrays were applied to profile plasma and serum samples from human DILI cases and controls. Methods: An initial screening was performed using 4594 randomly selected antibodies, representing 3450 human proteins. Resulting candidate proteins together with proposed DILI biomarker candidates generated a DILI array of 251 proteins for subsequent target analysis and verifications. In total, 1196 samples from 241 individuals across four independent cohorts were profiled: healthy volunteers receiving acetaminophen, patients with human immunodeficiency virus and/or tuberculosis receiving treatment, DILI cases originating from a wide spectrum of drugs, and healthy volunteers receiving heparins. Results: We observed elevated levels of cadherin 5, type 2 (CDH5) and fatty acid-binding protein 1 (FABP1) in DILI cases. In the two longitudinal cohorts, CDH5 was elevated already at baseline. FABP1 was elevated after treatment initiation and seemed to respond more rapidly than alanine aminotransferase (ALT). The elevations were verified in the DILI cases treated with various drugs. In the heparin cohort, CDH5 was stable over time whereas FABP1 was elevated. Conclusions: These results suggest that CDH5 may have value as a susceptibility marker for DILI. FABP1 was identified as a biomarker candidate with superior characteristics regarding tissue distribution and kinetics compared to ALT but likely with limited predictive value for the development of severe DILI. Further studies are needed to determine the clinical utility of the proposed markers. © 2016 John Wiley & Sons A/S.

  • 311. Mulder, J.
    et al.
    Björling, Erik
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Jonasson, Kalle
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Wernérus, Henrik
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Hober, Sophia
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Hokfelt, T.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Tissue Profiling of the Mammalian Central Nervous System Using Human Antibody-based Proteomics2009Inngår i: Molecular & Cellular Proteomics, ISSN 1535-9476, E-ISSN 1535-9484, Vol. 8, nr 7, s. 1612-1622Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    A need exists for mapping the protein profiles in the human brain both during normal and disease conditions. Here we studied 800 antibodies generated toward human proteins as part of a Human Protein Atlas program and investigated their suitability for detailed analysis of various levels of a rat brain using immuno-based methods. In this way, the parallel, rather limited analysis of the human brain, restricted to four brain areas (cerebellum, cerebral cortex, hippocampus, and lateral subventricular zone), could be extended in the rat model to 25 selected areas of the brain. Approximately 100 antibodies (12%) revealed a distinct staining pattern and passed validation of specificity using Western blot analysis. These antibodies were applied to coronal sections of the rat brain at 0.7-mm intervals covering the entire brain. We have now produced detailed protein distribution profiles for these antibodies and acquired over 640 images that form the basis of a publicly available portal of an antibody-based Rodent Brain Protein Atlas database (www.proteinatlas.org/rodentbrain). Because of the systematic selection of target genes, the majority of antibodies included in this database are generated against proteins that have not been studied in the brain before. Furthermore optimized tissue processing and colchicine treatment allow a high quality, more extended annotation and detailed analysis of subcellular distributions and protein dynamics. Molecular & Cellular Proteomics 8: 1612-1622, 2009.

  • 312. Mulder, J.
    et al.
    Wernérus, Henrik
    KTH, Skolan för bioteknologi (BIO), Molekylär Bioteknologi.
    Shi, T. J.
    Ponten, F.
    Hober, Sophia
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Hokfelt, T.
    Systematically generated antibodies against human gene products: High throughput screening on sections from the rat nervous system2007Inngår i: Neuroscience, ISSN 0306-4522, E-ISSN 1873-7544, Vol. 146, nr 4, s. 1689-1703Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Completion of the Human Genome Project and recent developments in proteomics make it possible to systematically generate affinity reagents to a large portion of the proteome. Recently an antibody-based human protein atlas covering many organs including four areas of the brain has been released (www.proteinatlas.org). Due to the heterogeneity, size, and availability of tissue a more thorough analysis of the human brain is associated with considerable difficulties. Here we applied 120 antibodies raised against 112 human gene products to the smaller rat brain, a rodent animal model, where a single section represents a 'superarray' including many brain areas, and consequently allowing analysis of a huge number of cell types and their neurochemicals. Immunoreactive structures were seen in the investigated brain tissue after incubation with 56 antibodies (46.6%), of which 25 (20.8%) showed a clearly discrete staining pattern that was limited to certain areas, or subsets of brain cells. Bioinformatics, pre-adsorption tests and Western blot analysis were applied to identify non-specific antibodies. Eleven antibodies, including such raised against four 'ambiguous' proteins, passed all validation criteria, and the expression pattern and subcellular distribution of these proteins were studied in detail. To further explore the potential of the systematically generated antibodies, all 11 antibodies that passed validation were used to analyze the spinal cord and lumbar dorsal root ganglia after unilateral transection of the sciatic nerve. Discrete staining patterns were observed for four of the proteins, and injury-induced regulation was found for one of them. In conclusion, the study presented here suggests that a significant portion (10%) of the antibodies generated to a human protein can be used to analyze orthologues present in the rodent brain and to produce a protein-based atlas of the rodent brain. It is hoped that this type of antibody-based, high throughput screening of brain tissue from various rodent disease models will provide new information on the brain chemical neuroanatomy and insights in processes underlying neurological pathologies.

  • 313. Mulder, Jan
    et al.
    Spence, Lauren
    Tortoriello, Giuseppe
    DiNieri, Jennifer A.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101).
    Shui, Bo
    Kotlikoff, Michael I.
    Yanagawa, Yuchio
    Aujard, Fabienne
    Hokfelt, Tomas
    Hurd, Yasmin L.
    Harkany, Tibor
    Secretagogin is a Ca2+-binding protein identifying prospective extended amygdala neurons in the developing mammalian telencephalon2010Inngår i: European Journal of Neuroscience, ISSN 0953-816X, E-ISSN 1460-9568, Vol. 31, nr 12, s. 2166-2177Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The Ca2+-binding proteins (CBPs) calbindin D28k, calretinin and parvalbumin are phenotypic markers of functionally diverse subclasses of neurons in the adult brain. The developmental dynamics of CBP expression are precisely timed: calbindin and calretinin are present in prospective cortical interneurons from mid-gestation, while parvalbumin only becomes expressed during the early postnatal period in rodents. Secretagogin (scgn) is a CBP cloned from pancreatic beta and neuroendocrine cells. We hypothesized that scgn may be expressed by particular neuronal contingents during prenatal development of the mammalian telencephalon. We find that scgn is expressed in neurons transiting in the subpallial differentiation zone by embryonic day (E)11 in mouse. From E12, scgn+ cells commute towards the extended amygdala and colonize the bed nucleus of stria terminalis, the interstitial nucleus of the posterior limb of the anterior commissure, the dorsal substantia innominata (SI) and the central and medial amygdaloid nuclei. Scgn+ neurons can acquire a cholinergic phenotype in the SI or differentiate into GABA cells in the central amygdala. We also uncover phylogenetic differences in scgn expression as this CBP defines not only neurons destined to the extended amygdala but also cholinergic projection cells and cortical pyramidal cells in the fetal nonhuman primate and human brains, respectively. Overall, our findings emphasize the developmentally shared origins of neurons populating the extended amygdala, and suggest that secretagogin can be relevant to the generation of functional modalities in specific neuronal circuitries.

  • 314. Mulder, Jan
    et al.
    Zilberter, Misha
    Spence, Lauren
    Tortoriello, Giuseppe
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101).
    Yanagawa, Yuchio
    Aujard, Fabienne
    Hokfelt, Tomas
    Harkany, Tibor
    Secretagogin is a Ca2+-binding protein specifying subpopulations of telencephalic neurons2009Inngår i: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 106, nr 52, s. 22492-22497Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The Ca2+-binding proteins (CBPs) parvalbumin, calbindin, and calretinin are phenotypic markers of terminally differentiated neurons in the adult brain. Although subtle phylogenetic variations in the neuronal distribution of these CBPs may occur, morphologically and functionally diverse subclasses of interneurons harbor these proteins in olfactory and corticolimbic areas. Secretagogin (scgn) is a recently cloned CBP from pancreatic beta and neuroendocrine cells. We hypothesized that scgn is expressed in the mammalian brain. We find that scgn is a marker of neuroblasts commuting in the rostral migratory stream. Terminally differentiated neurons in the olfactory bulb retain scgn expression, with scgn being present in periglomerular cells and granular layer interneurons. In the corticolimbic system, scgn identifies granule cells distributed along the dentate gyrus, indusium griseum, and anterior hippocampal continuation emphasizing the shared developmental origins, and cytoarchitectural and functional similarities of these neurons. We also uncover unexpected phylogenetic differences in scgn expression, since this CBP is restricted to primate cholinergic basal forebrain neurons. Overall, we characterize scgn as a neuron-specific CBP whose distribution identifies neuronal subtypes and hierarchical organizing principles in the mammalian brain.

  • 315.
    Neiman, Maja
    et al.
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Fredolini, Claudia
    KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Johansson, H.
    Lehtiö, Janne
    Karolinska Institutet, Solna, Sweden .
    Nygren, Per-Åke
    KTH, Skolan för bioteknologi (BIO), Proteinteknologi.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Nilsson, Peter
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Schwenk, Jochen M.
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Selectivity analysis of single binder assays used in plasma protein profiling2013Inngår i: Proteomics, ISSN 1615-9853, E-ISSN 1615-9861, Vol. 13, nr 23-24, s. 3406-3410Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The increasing availability of antibodies toward human proteins enables broad explorations of the proteomic landscape in cells, tissues, and body fluids. This includes assays with antibody suspension bead arrays that generate protein profiles of plasma samples by flow cytometer analysis. However, antibody selectivity is context dependent so it is necessary to corroborate on-target detection over off-target binding. To address this, we describe a concept to directly verify interactions from antibody-coupled beads by analysis of their eluates by Western blots and MS. We demonstrate selective antibody binding in complex samples with antibodies toward a set of chosen proteins with different abundance in plasma and serum, and illustrate the need to adjust sample and bead concentrations accordingly. The presented approach will serve as an important tool for resolving differential protein profiles from antibody arrays within plasma biomarker discoveries.

  • 316.
    Neiman, Maja
    et al.
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Hedberg, Jesper J.
    Dönnes, Pierre R.
    Schuppe-Koistinen, Ina
    Hanschke, Stephan
    Schindler, Ralf
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Schwenk, Jochen M.
    KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Nilsson, Peter
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Plasma Profiling Reveals Human Fibulin-1 as Candidate Marker for Renal Impairment2011Inngår i: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 10, nr 11, s. 4925-4934Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    There is a need for reliable and sensitive biomarkers for renal impairments to detect early signs of kidney toxicity and to monitor progression of disease. Here, antibody suspension bead arrays were applied to profile plasma samples from patients with four types of kidney disorders: glomerulonephritis, diabetic nephropathy, obstructive uropathy, and analgesic abuse. In total, 200 clinical renal-associated cases and control plasma samples from different cohorts were profiled. Parallel plasma protein profiles were obtained using biotinylated and nonfractionated samples and a selected set of 94 proteins targeted by 129 antigen-purified polyclonal antibodies. Out of the analyzed target proteins, human fibulin-1 was detected at significantly higher levels in the glomerulonephritis patient group compared to the controls and with elevated levels in patient samples for all other renal disorders investigated. Two polyclonal antibodies and one monoclonal antibody directed toward separate, nonoverlapping epitopes showed the same trend in the discovery cohorts. A technical verification using Western blot analysis of selected patient plasma confirmed the trends toward higher abundance of the target protein in disease samples. Furthermore, a verification study was carried out in the context of glomerulonephritis using an independent case and control cohort, and this confirmed the results from the discovery cohort, suggesting that plasma levels of fibulin-1 could serve as a potential indicator to monitor kidney malfunction or kidney damage.

  • 317.
    Neiman, Maja
    et al.
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap, Affinity Proteomics.
    Hellström, Cecilia
    KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap, Affinity Proteomics. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Just, David
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap, Affinity Proteomics.
    Mattsson, Cecilia
    KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap, Affinity Proteomics. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Fagerberg, Linn
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap, Systembiologi.
    Schuppe-Koistinen, Ina
    KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap, Systembiologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Gummesson, Anders
    Sahlgrens Univ Hosp, Dept Clin Pathol & Genet, Gothenburg, Sweden..
    Bergstrom, Goran
    Sahlgrens Univ Hosp, Dept Clin Physiol, Gothenburg, Sweden..
    Kallioniemi, Olli
    Univ Helsinki, Inst Mol Med Finland, FIMM, Helsinki, Finland.;Karolinska Inst, Dept Pathol & Oncol, SciLifeLab, Stockholm, Sweden..
    Achour, Adnane
    Karolinska Inst, SciLifeLab, Dept Med Solna, Stockholm, Sweden.;Karolinska Univ Hosp, Div Infect Dis, Stockholm, Sweden..
    Sallinen, Riitta
    Univ Helsinki, Inst Mol Med Finland, FIMM, Helsinki, Finland.;Karolinska Inst, Dept Pathol & Oncol, SciLifeLab, Stockholm, Sweden..
    Uhlén, Mathias
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap, Systembiologi.
    Nilsson, Peter
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap, Affinity Proteomics.
    Individual and stable autoantibody repertoires in healthy individuals2019Inngår i: Autoimmunity, ISSN 0891-6934, E-ISSN 1607-842X, Vol. 52, nr 1, s. 1-11Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    In the era towards precision medicine, we here present the individual specific autoantibody signatures of 193 healthy individuals. The self-reactive IgG signatures are stable over time in a way that each individual profile is recognized in longitudinal sampling. The IgG autoantibody reactivity towards an antigen array comprising 335 protein fragments, representing 204 human proteins with potential relevance to autoimmune disorders, was measured in longitudinal plasma samples from 193 healthy individuals. This analysis resulted in unique autoantibody barcodes for each individual that were maintained over one year's time. The reactivity profiles, or signatures, are person specific in regards to the number of reactivities and antigen specificity. Two independent data sets were consistent in that each healthy individual displayed reactivity towards 0-16 antigens, with a median of six. Subsequently, four selected individuals were profiled on in-house produced high-density protein arrays containing 23,000 protein fragments representing 14,000 unique protein coding genes. Based on a unique, broad and deep longitudinal profiling of autoantibody reactivities, our results demonstrate a unique autoreactive profile in each analyzed healthy individual. The need and interest for broad-ranged and high-resolution molecular profiling of healthy individuals is rising. We have here generated and assessed an initial perspective on the global distribution of the self-reactive IgG repertoire in healthy individuals, by investigating 193 well-characterized healthy individuals.

  • 318.
    Neiman, Maja
    et al.
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Nygren, Per-Åke
    KTH, Skolan för bioteknologi (BIO), Centra, Albanova VinnExcellence Center for Protein Technology, ProNova. KTH, Skolan för bioteknologi (BIO), Molekylär Bioteknologi (stängd 20130101).
    Uhlén, Mathias
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101).
    Nilsson, Peter
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Schwenk, Jochen M.
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Validating the selectivity of antibodies used in multiplexed serum profiling via parallel immunocapture analysisManuskript (preprint) (Annet vitenskapelig)
    Abstract [en]

    The increasing availability of antibodies towards human proteins drives the generation of new and exploratory data, which can be generated by profiling e.g. plasma samples using multiplexed arrays. However, antibody assays can lead to erroneous results due to cross-­‐reactivity to off-­‐targets. Here, we describe an approach in which an antibody-­‐based suspension bead array is combined with subsequent validation of on-­‐ target binding using a coupled affinity purification assay. Based on differential heat treatment of samples, antibody performance was investigated and the results for antibodies directed towards several complement factors provide insights on the detection of proteins in sera from patients with complement deficiency. In conclusion, the combined parallel flow and blot based immunocapture analysis serves an important, first line tool for resolving differently detected serum or plasma protein profiles proposed by antibody arrays. 

  • 319. Newberg, J. Y.
    et al.
    Li, J.
    Rao, A.
    Pontén, F.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101).
    Lundberg, Emma
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101).
    Murphy, R. F.
    Automated analysis of human protein atlas immunofluorescence images2009Inngår i: Proceedings - 2009 IEEE International Symposium on Biomedical Imaging: From Nano to Macro, ISBI 2009, IEEE , 2009, s. 1023-1026Konferansepaper (Fagfellevurdert)
    Abstract [en]

    The Human Protein Atlas is a rich source of location proteomics data. In this work, we present an automated approach for processing and classifying major subcellular patterns in the Atlas images. We demonstrate that two different classification frameworks (support vector machine and random forest) are effective at determining subcellular locations; we can analyze over 3500 Atlas images with a high degree of accuracy, up to 87.5% for all of the samples and 98.5% when only considering samples in whose classification assignments we are most confident. Moreover, the features obtained in both of these frameworks are observed to be highly consistent and generalizable. Additionally, we observe that the features relating the proteins to cell markers are especially important in automated learning approaches.

  • 320. Nguyen, N
    et al.
    Samuelson, Patrik
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Sterky, Fredrik
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Merle-Poitte, C
    Robert, A
    Baussant, T
    Haeuw, F
    Uhlen, Mathias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Binz, H
    Ståhl, Stefan
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Chromosomal sequencing using a PCR-based biotin-capture method allowed isolation of the complete gene for the outer membrane protein A of Klebsiella pneumoniae1998Inngår i: Gene, ISSN 0378-1119, E-ISSN 1879-0038, Vol. 210, nr 1, s. 93-101Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    By employing a novel biotin-and PCR-assisted capture method, which allows determination of unknown sequences on chromosomal DNA. the gene for the outer membrane protein A (OmpA) of Klebsiella pneumoniae has been isolated and sequenced to completion. The method involves linear amplification of DNA from a biotinylated primer annealing to a region with known sequence. After capture of the amplified single-stranded DNA on to paramagnetic beads, unspecifically annealing primers, i.e. arbitrary primers, were used to generate fragments with only partly determined nt sequences. The homology of the sequenced gene to ompAs of related bacteria is discussed. The ompA gene was assembled for intracellular expression in Escherichia coli, and two different fusion proteins were produced and recovered with good yields. The importance of the novel chromosomal sequencing method for gene isolation in general and the potential use of the OmpA fusion proteins are discussed. (C) 1998 Elsevier Science B.V.

  • 321. Nilsson, Björn
    et al.
    Nygren, Per-Åke
    KTH, Tidigare Institutioner, Bioteknologi.
    Uhlén, Mathias
    KTH, Tidigare Institutioner, Bioteknologi.
    Bacterial receptor structures1994Patent (Annet (populærvitenskap, debatt, mm))
    Abstract [en]

    Novel proteins obtainable by mutagenesis of surface-exposed amino acids of domains of natural bacterial receptors, said proteins being obtained without substantial loss of basic structure and stability of said natural bacterial receptors; proteins which have been selected from a protein library embodying a repertoire of said novel proteins; and methods for the manufacture of artificial bacterial receptor structures.

  • 322. Nilsson, Carol L.
    et al.
    Mostovenko, Ekaterina
    Lichti, Cheryl F.
    Ruggles, Kelly
    Fenyoe, David
    Rosenbloom, Kate R.
    Hancock, William S.
    Paik, Young-Ki
    Omenn, Gilbert S.
    LaBaer, Joshua
    Kroes, Roger A.
    Uhlén, Matthias
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Hober, Sophia
    KTH, Skolan för bioteknologi (BIO), Proteinteknologi.
    Vegvari, Akos
    Andren, Per E.
    Sulman, Erik P.
    Lang, Frederick F.
    Fuentes, Manuel
    Carlsohn, Elisabet
    Emmett, Mark R.
    Moskal, Joseph R.
    Berven, Frode S.
    Fehniger, Thomas E.
    Marko-Varga, Gyorgy
    Use of ENCODE Resources to Characterize Novel Proteoforms and Missing Proteins in the Human Proteome2015Inngår i: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 14, nr 2, s. 603-608Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    We describe the utility of integrated strategies that employ both translation of ENCODE data and major proteomic technology pillars to improve the identification of the "missing proteins", novel proteoforms, and PTMs. On one hand, databases in combination with bioinformatic tools are efficiently utilized to establish microarray-based transcript analysis and supply rapid protein identifications in clinical samples. On the other hand, sequence libraries are the foundation of targeted protein identification and quantification using mass spectrometric and immunoaffinity techniques. The results from combining proteoENCODEdb searches with experimental mass spectral data indicate that some alternative splicing forms detected at the transcript level are in fact translated to proteins. Our results provide a step toward the directives of the C-HPP initiative and related biomedical research.

  • 323.
    Nilsson, P.
    et al.
    KTH.
    Janzi, M.
    KTH.
    Odling, J.
    KTH.
    Sundberg, M.
    KTH.
    Lundeberg, Joakim
    KTH.
    Uhlén, Mathias
    KTH, Tidigare Institutioner (före 2005), Bioteknologi.
    Hammarstrom, L.
    KTH.
    Serum microarrays for screening of protein levels2005Inngår i: Molecular & Cellular Proteomics, ISSN 1535-9476, E-ISSN 1535-9484, Vol. 4, nr 8, s. S334-S334Artikkel i tidsskrift (Annet vitenskapelig)
  • 324.
    Nilsson, Peter
    et al.
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Larsson, A
    Lundeberg, Joakim
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Uhlen, Mathias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Nygren, Per-Åke
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Mutational scanning of PCR products by subtractive oligonucleotide hybridization analysis1999Inngår i: BioTechniques, ISSN 0736-6205, E-ISSN 1940-9818, Vol. 26, nr 2, s. 308-+Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Here, we describe a new approach for mutational scanning of PCR products through hybridization analysis between complementary oligonucleotides. Sets of overlapping probe oligonucleotides complementary to wild-type (WT) sequence are hybridized to microbead-immobilized PCR products under solution-like conditions. Mismatch-hybridization situations between a mutant sample and probe oligonucleotides result in higher remaining concentrations in solution of involved probe oligonucleotides. Post-hybridization supernatants are subsequently analyzed for their probe oligonucleotide compositions using surface plasmon resonance-based biosensor technology Relative remaining probe oligonucleotide concentrations are monitored in real-time through hybridization analysis between probe oligonucleotides and their corresponding sensor-chip immobilized complementary counterparts. This allows for the construction of composition diagrams revealing the existence and approximate location of a mutation within an investigated sample DNA sequence. Applied on PCR products derived from clinical samples of microdissected tumor biopsies, single mutations in exons 6 and 7 of the human p53 tumor-suppressor gene were successfully detected and approximately localized.

  • 325.
    Nilsson, Peter
    et al.
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Larsson, Karin
    KTH, Skolan för bioteknologi (BIO).
    Persson, Anja
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Wernérus, Henrik
    KTH, Skolan för bioteknologi (BIO).
    Hober, Sophia
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Björling, Erik
    KTH, Skolan för bioteknologi (BIO).
    Ottoson, Jenny
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Ödling, Jenny
    KTH, Skolan för bioteknologi (BIO).
    Sundberg, Mårten
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Al-Khalili Szigyarto, Cristina
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Paavilainen, Linda
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden.
    Andersson, Ann-Catrin
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden.
    Kampf, Caroline
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden.
    Wester, Kenneth
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden.
    Pontén, Fredrik
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden.
    Towards a human proteome atlas: High-throughput generation of mono-specific antibodies for tissue profiling2005Inngår i: Proteomics, ISSN 1615-9853, E-ISSN 1615-9861, Vol. 5, s. 4327-4337Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    A great need exists for the systematic generation of specific antibodies to explore the human proteome. Here, we show that antibodies specific to human proteins can be generated in a high-throughput manner involving stringent affinity purification using recombinant protein epitope signature tags (PrESTs) as immunogens and affinity-ligands. The specificity of the generated affinity reagents, here called mono-specific antibodies (msAb), were validated with a novel protein microarray assay. The success rate for 464 antibodies generated towards human proteins was more than 90% as judged by the protein array assay. The antibodies were used for parallel profiling of patient biopsies using tissue microarrays generated from 48 human tissues. Comparative analysis with well-characterized monoclonal antibodies showed identical or similar specificity and expression patterns. The results suggest that a comprehensive atlas containing extensive protein expression and subcellular localization data of the human proteome can be generated in an efficient manner with mono-specific antibodies.

  • 326.
    Nilsson, Peter
    et al.
    KTH, Tidigare Institutioner                               , Bioteknologi.
    O'Meara, Deirdre
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Edebratt, Fredrik
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Persson, Björn
    Uhlén, Mattias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Lundeberg, Joakim
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Nygren, Per-Åke
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Quantitative Investigation of the Modular Primer Effect for DNA and Peptide Nucleic Acid Hexamers1999Inngår i: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 269, nr 1, s. 155-161Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The effect on oligonucleotide–template duplex stability upon cohybridization of adjacently annealing oligonucleotides, the modular primer effect, was studied with biosensor technology. DNA and peptide nucleic acid (PNA) hexamer modules and sensor chip-immobilized template DNA strands were designed for analysis of nick, overlap, and gap modular hybridization situations. The fast hybridization kinetics for such hexamer modules allowed for the determination of apparent duplex affinities from equilibrium responses. The results showed that the hybridizational stability of modular hexamer pairs is strongly dependent on the positioning, concentration, and inherent affinity of the adjacently annealing hexamer module. Up to 80-fold increases in apparent affinities could be observed for adjacent modular oligonucleotide pairs compared to affinities determined for single hexamer oligonucleotide hybridizations. Interestingly, also for coinjections of different module combinations where DNA hexamer modules were replaced by their PNA counterparts, a modular primer effect was observed. The introduction of a single base gap between two hexamer modules significantly reduced the stabilization effect, whereas a gap of two bases resulted in a complete loss of the effect. The results suggest that the described biosensor-based methodology should be useful for the selection of appropriate modules and working concentrations for use in different modular hybridization applications.

  • 327.
    Nilsson, Peter
    et al.
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Persson, B
    Larsson, A
    Uhlen, Mathias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Nygren, Per-Åke
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Detection of mutations in PCR products from clinical samples by surface plasmon resonance1997Inngår i: Journal of Molecular Recognition, ISSN 0952-3499, E-ISSN 1099-1352, Vol. 10, nr 1, s. 7-17Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Two different strategies for scanning and screening of mutations in polymerase chain reaction (PCR) products by hybridization analysis are described, employing real-time biospecific interaction analysis (BIA) for detection, Real-time BIA was used to detect differences in hybridization responses between PCR products and different 17-mer oligonucleotide probes, For the analysis using a biosensor instrument, two different experimental formats were investigated based on immobilization of either biotinylated PCR products or oligonucleotide probes onto a sensor chip, Applied on the human tumour suppressor p53 gene, differences in hybridization levels for full-match and mismatch situations employing both formals allowed the detection of point mutations in exon 6 PCR products, derived from a breast tumour biopsy sample, In addition, a mutant sample sequence could be detected in a 50/50 background of wild type exon 6 sequence, The suitability of the different formats for obtaining a regenerable system and a high throughput of samples is discussed.

  • 328.
    NILSSON, PETER
    et al.
    KTH, Tidigare Institutioner, Bioteknologi.
    PERSSON, BJÖRN
    UHLEN, MATHIAS
    KTH, Tidigare Institutioner, Bioteknologi.
    NYGREN, PER-ÅKE
    KTH, Tidigare Institutioner, Bioteknologi.
    REAL-TIME MONITORING OF DNA MANIPULATIONS USING BIOSENSOR TECHNOLOGY1995Inngår i: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 224, nr 1, s. 400-408Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The potential of real-time biospecific interaction analysis technology for applications in molecular biology is described. DNA fragments are immobilized onto a biosensor surface using the high-affinity streptavidin-biotin system and subsequently used to monitor different unit operations in molecular biology, e.g., DNA strand separation, DNA hybridization kinetics, and enzymatic modifications. A model system comprising six oligonucleotides was used, which can be assembled into a 69-bp double-stranded DNA fragment. Using this system, the biosensor approach was employed to analyze multistep solid-phase gene assembly and the performance of different enzymes routinely used for the synthesis and manipulation of DNA. In addition, a concept for the determination of single-point mutations in DNA samples is described.

  • 329.
    Nilsson, Peter
    et al.
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Schwenk, Jochen
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Biomarker of renal impairment2011Patent (Annet (populærvitenskap, debatt, mm))
    Abstract [en]

    There is provided a method of determining whether a subject belongs to a first or a second group of subjects, wherein the risk of having or developing of a renal impairment is higher in the first group than in the second group, comprising the steps of: a) measuring an amount of fibulin 1 in a sample from the subject to obtain a sample value; b) comparing the sample value to a reference value; and if the sample value is higher than the reference value, c1) concluding that the subject belongs to the first group; and if the sample value is lower than the previous sample value, c2) concluding that the subject belongs to the second group. Associated means are also provided.

  • 330.
    Nilsson, Peter
    et al.
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Schwenk, Jochen
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Prostate cancer biomarkers2010Patent (Annet (populærvitenskap, debatt, mm))
    Abstract [en]

    The present disclosure relates to a method of determining whether a mammalian subject belongs to a first or a second group of subjects, wherein the risk of having a prostate cancer, such as an aggressive prostate cancer, is higher in the first group than in the second group. The method comprises the steps of: a) evaluating an amount of unglycosylated CNDP1 protein in a sample derived from blood, semen or urine from the subject and determining a sample value corresponding to the evaluated amount; b) comparing the sample value with a predetermined reference value; and if the sample value is lower than or equal to the reference value, c1) concluding that the subject belongs to the first group; and if the sample value is higher than the reference value, c2) concluding that the subject belongs to the second group. Further, the present disclosure relates to a similar method based on the protein HCRTR1 as well as corresponding uses and means useful when carrying out the methods.

  • 331. Nishibori, Yukino
    et al.
    Katayama, Kan
    Parikka, Mataleena
    Oddsson, Asmundur
    Nukui, Masatoshi
    Hultenby, Kjell
    Wernerson, Annika
    He, Bing
    Ebarasi, Lwaki
    Raschperger, Elisabeth
    Norlin, Jenny
    Uhlen, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Patrakka, Jaakko
    Betsholtz, Christer
    Tryggvason, Karl
    Glcci1 Deficiency Leads to Proteinuria2011Inngår i: Journal of the American Society of Nephrology, ISSN 1046-6673, E-ISSN 1533-3450, Vol. 22, nr 11, s. 2037-2046Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Unbiased transcriptome profiling and functional genomics approaches identified glucocorticoid-induced transcript 1 (GLCCI1) as being a transcript highly specific for the glomerulus, but its role in glomerular development and disease is unknown. Here, we report that mouse glomeruli express far greater amounts of Glcci1 protein compared with the rest of the kidney. RT-PCR and Western blotting demonstrated that mouse glomerular Glcci1 is approximately 60 kD and localizes to the cytoplasm of podocytes in mature glomeruli. In the fetal kidney, intense Glcci1 expression occurs at the capillary-loop stage of glomerular development. Using gene knockdown in zebrafish with morpholinos, morphants lacking Glcci1 function had collapsed glomeruli with foot-process effacement. Permeability studies of the glomerular filtration barrier in these zebrafish morphants demonstrated a disruption of the selective glomerular permeability filter. Taken together, these data suggest that Glcci1 promotes the normal development and maintenance of podocyte structure and function.

  • 332. Nodin, Bjorn
    et al.
    Fridberg, Marie
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Jirström, Karin
    Discovery of Dachshund 2 protein as a novel biomarker of poor prognosis in epithelial ovarian cancer2012Inngår i: Journal of Ovarian Research, ISSN 1757-2215, Vol. 5, nr 1, s. 6-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: The Dachshund homolog 2 (DACH2) gene has been implicated in development of the female genital tract in mouse models and premature ovarian failure syndrome, but to date, its expression in human normal and cancerous tissue remains unexplored. Using the Human Protein Atlas as a tool for cancer biomarker discovery, DACH2 protein was found to be differentially expressed in epithelial ovarian cancer (EOC). Here, the expression and prognostic significance of DACH2 was further evaluated in ovarian cancer cell lines and human EOC samples. Methods: Immunohistochemical expression of DACH2 was examined in tissue microarrays with 143 incident EOC cases from two prospective, population-based cohorts, including a subset of benign-appearing fallopian tubes (n = 32). A nuclear score (NS), i.e. multiplier of staining fraction and intensity, was calculated. For survival analyses, cases were dichotomized into low (NS < = 3) and high (NS > 3) using classification and regression tree analysis. Kaplan Meier analysis and Cox proportional hazards modelling were used to assess the impact of DACH2 expression on survival. DACH2 expression was analysed in the cisplatin sensitive ovarian cancer cell line A2780 and its cisplatin resistant derivative A2780-Cp70. The specificity of the DACH2 antibody was tested using siRNA-mediated silencing of DACH2 in A2780-Cp70 cells. Results: DACH2 expression was considerably higher in the cisplatin resistant A2780-Cp70 cells compared to the cisplatin-sensitive A2780 cells. While present in all sampled fallopian tubes, DACH2 expression ranged from negative to strong in EOC. In EOC, DACH2 expression correlated with several proteins involved in DNA integrity and repair, and proliferation. DACH2 expression was significantly higher in carcinoma of the serous subtype compared to non-serous carcinoma. In the full cohort, high DACH2 expression was significantly associated with poor prognosis in univariable analysis, and in carcinoma of the serous subtype, DACH2 remained an independent factor of poor prognosis. Conclusions: This study provides a first demonstration of DACH2 protein being expressed in human fallopian tubes and EOC, with the highest expression in serous carcinoma where DACH2 was found to be an independent biomarker of poor prognosis. Future research should expand on the role of DACH2 in ovarian carcinogenesis and chemotherapy resistance.

  • 333. Nodin, Björn
    et al.
    Fridberg, Marie
    Jonsson, Liv
    Bergman, Julia
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Jirström, Karin
    High MCM3 expression is an independent biomarker of poor prognosis and correlates with reduced RBM3 expression in a prospective cohort of malignant melanoma2012Inngår i: Diagnostic Pathology, ISSN 1746-1596, E-ISSN 1746-1596, Vol. 7, nr 1, s. 82-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: Malignant melanoma is the most lethal form of skin cancer with a variable clinical course even in patients with thin melanomas and localized disease. Despite increasing insights into melanoma biology, no prognostic biomarkers have yet been incorporated into clinical protocols. Reduced expression of the RNA binding motif protein 3 (RBM3) has been shown to correlate with tumour progression and poor prognosis in melanoma and several other cancer forms. In ovarian cancer, an inverse association was found between expression of RBM3 and the minichromosome maintenance 3 (MCM3) gene and protein. In melanoma, gene expression analysis and immunohistochemical validation has uncovered MCM3 as a putative prognostic biomarker. The aim of the present study was to examine the associations of MCM3 expression with clinical outcome and RBM3 expression in a prospective, population-based cohort of melanoma. Methods: Immunohistochemical MCM3 expression was examined in 224 incident cases of primary melanoma from the Malmo Diet and Cancer Study, previously analysed for RBM3 expression. Spearman's Rho and Chi-Square tests were used to explore correlations between MCM3 expression, clinicopathological factors, and expression of RBM3 and Ki67. Kaplan Meier analysis, the log rank test, and univariable and multivariable Cox proportional hazards modelling were used to assess the impact of MCM3 expression on disease-free survival (DFS) and melanoma-specific survival (MSS). Results: High MCM3 expression was significantly associated with unfavourable clinicopathological features and high Ki67 expression. A significant inverse correlation was seen between expression of MCM3 and RBM3 (p = 0.025). High MCM3 expression was associated with a reduced DFS (HR = 5.62) and MSS (HR = 6.03), and these associations remained significant in multivariable analysis, adjusted for all other factors (HR = 5.01 for DFS and HR = 4.96 for MSS). RBM3 expression remained an independent prognostic factor for MSS but not DFS in the multivariable model. Conclusions: These findings provide validation of the utility of MCM3 expression as an independent biomarker for prognostication of patients with primary melanoma. Moreover, the inverse association and prognostic impact of MCM3 and RBM3 expression indicate a possible interaction of these proteins in melanoma progression, the functional basis for which merits further study.

  • 334. Nodin, Björn
    et al.
    Hedner, Charlotta
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Fridberg, Marie
    Jirström, Karin
    Expression of the global regulator SATB1 is an independent factor of poor prognosis in high grade epithelial ovarian cancer2012Inngår i: Tumor Biology, ISSN 1010-4283, E-ISSN 1423-0380, Vol. 33, s. 89-89Artikkel i tidsskrift (Annet vitenskapelig)
  • 335. Nodin, Björn
    et al.
    Hedner, Charlotta
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Jirström, Karin
    Expression of the global regulator SATB1 is an independent factor of poor prognosis in high grade epithelial ovarian cancer2012Inngår i: Journal of Ovarian Research, ISSN 1757-2215, Vol. 5, nr 1, s. 24-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: The global gene regulator Special AT-rich sequence-binding protein1 (SATB1) has been reported to reprogramme tumour cells into a more malignant phenotype and associate with poor clinical outcome in several cancer forms. In this study, we investigated the molecular correlates and prognostic impact of SATB1 expression in human epithelial ovarian cancer (EOC). Findings: Immunohistochemical expression of SATB1 was examined in tissue microarrays with tumours from 151 incident EOC cases from two prospective, population-based cohorts. Benign-appearing fallopian tube epithelium from 32 cases was also analyzed. A multiplier of nuclear fraction and staining intensity of SATB1 was calculated. While barely expressed in tubal epithelium, nuclear SATB1 expression was denoted in 35/151 (23.2%) EOC cases. Spearman's Rho test revealed an inverse correlation between SATB1 expression and histological grade (R = -0.22, p = 0.006) and a positive correlation with expression of dachshund 2 protein (R = 0.28, p = 0.001), phosphorylated Chek1 (R = 0.26, p = 0.002) and minichromosome maintenance protein 3 (R = 0.17, p = 0.042). Univariable Cox regression analysis revealed that SATB1 expression, while not prognostic in the full cohort, was associated with a reduced ovarian cancer-specific survival and 5-year overall survival in high grade tumours (n = 105) (HR = 2.14 and HR = 1.96, respectively). This association remained significant in multivariable analysis, adjusted for age and clinical stage (HR = 2.20 and HR = 2.06, respectively). Conclusions: These results demonstrate that SATB1 expression is an independent factor of poor prognosis in high grade EOC and correlates in vivo with cellular processes involved in the maintenance of DNA integrity. The functional basis for these observations merits further investigation.

  • 336. Nodin, Björn
    et al.
    Johannesson, Henrik
    Wangefjord, Sakarias
    O'Connor, Darran P.
    Lindquist, Kajsa Ericson
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101). KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Jirström, Karin
    Eberhard, Jakob
    Molecular correlates and prognostic significance of SATB1 expression in colorectal cancer2012Inngår i: Diagnostic Pathology, ISSN 1746-1596, E-ISSN 1746-1596, Vol. 7, nr 1, s. 115-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: Special AT-rich sequence-binding protein 1 (SATB1) is a global gene regulator that has been reported to confer malignant behavior and associate with poor prognosis in several cancer forms. SATB1 expression has been demonstrated to correlate with unfavourable tumour characteristics in rectal cancer, but its association with clinical outcome in colorectal cancer (CRC) remains unclear. In this study, we examined the prognostic impact of SATB1 expression in CRC, and its association with important molecular characteristics; i.e. beta-catenin overexpression, microsatellite instability (MSI) screening status, and SATB2 expression. Methods: Immunohistochemical expression of SATB1 and beta-catenin was assessed in tissue microarrays with tumours from 529 incident CRC cases in the prospective population-based Malmo Diet and Cancer Study, previously analysed for SATB2 expression and MSI screening status. Spearman's Rho and Chi-Square tests were used to explore correlations between SATB1 expression, clinicopathological and investigative parameters. Kaplan Meier analysis and Cox proportional hazards modelling were used to explore the impact of SATB1 expression on cancer specific survival (CSS) and overall survival (OS). Results: SATB1 was expressed in 222 (42%) CRC cases and negative, or sparsely expressed, in adjacent colorectal mucosa (n = 16). SATB1 expression was significantly associated with microsatellite stable tumours (p < 0.001), beta-catenin overexpression (p < 0.001) and SATB2 expression (p < 0.001). While not prognostic in the full cohort, SATB1 expression was significantly associated with poor prognosis in SATB2 negative tumours (HR = 2.63; 95% CI 1.46-4.71; p(interaction) = 0.011 for CSS and HR = 2.31; 95% CI 1.32-4.04; p(interaction) = 0.015 for OS), remaining significant in multivariable analysis. Conclusions: The results of this study demonstrate that SATB1 expression in CRC is significantly associated with beta-catenin overexpression, microsatellite stability and SATB2 expression. Furthermore, SATB1 expression is a factor of poor prognosis in SATB2 negative tumours. Altogether, these data indicate an important role for SATB1 in colorectal carcinogenesis and suggest prognostically antagonistic effects of SATB1 and SATB2. The mechanistic basis for these observations warrants further study.

  • 337. Nookaew, Intawat
    et al.
    Papini, Marta
    Pornputtapong, Natapol
    Scalcinati, Gionata
    Fagerberg, Linn
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Uhlén, Matthias
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Nielsen, Jens
    A comprehensive comparison of RNA-Seq-based transcriptome analysis from reads to differential gene expression and cross-comparison with microarrays: a case study in Saccharomyces cerevisiae2012Inngår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 40, nr 20, s. 10084-10097Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    RNA-seq, has recently become an attractive method of choice in the studies of transcriptomes, promising several advantages compared with microarrays. In this study, we sought to assess the contribution of the different analytical steps involved in the analysis of RNA-seq data generated with the Illumina platform, and to perform a cross-platform comparison based on the results obtained through Affymetrix microarray. As a case study for our work we, used the Saccharomyces cerevisiae strain CEN.PK 113-7D, grown under two different conditions (batch and chemostat). Here, we asses the influence of genetic variation on the estimation of gene expression level using three different aligners for read-mapping (Gsnap, Stampy and TopHat) on S288c genome, the capabilities of five different statistical methods to detect differential gene expression (baySeq, Cuffdiff, DESeq, edgeR and NOISeq) and we explored the consistency between RNA-seq analysis using reference genome and de novo assembly approach. High reproducibility among biological replicates (correlation >= 0.99) and high consistency between the two platforms for analysis of gene expression levels (correlation >= 0.91) are reported. The results from differential gene expression identification derived from the different statistical methods, as well as their integrated analysis results based on gene ontology annotation are in good agreement. Overall, our study provides a useful and comprehensive comparison between the two platforms (RNA-seq and microrrays) for gene expression analysis and addresses the contribution of the different steps involved in the analysis of RNA-seq data.

  • 338. Nord, K.
    et al.
    Gunneriusson, E.
    Uhlén, Mathias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Nygren, Per-Åke
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Ligands selected from combinatorial libraries of protein A for use in affinity capture of apolipoprotein A-1(M) and Taq DNA polymerase2000Inngår i: Journal of Biotechnology, ISSN 0168-1656, E-ISSN 1873-4863, Vol. 80, nr 1, s. 45-54Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Here we show that robust and small protein ligands can be used for affinity capture of recombinant proteins from crude cell lysates. Two ligands selectively binding to bacterial Tag DNA polymerase and human apolipoprotein A-1(M), respectively, were used in the study. The ligands were selected from libraries of a randomized alpha-helical bacterial receptor domain derived from staphylococcal protein A and have dissociation constants in the micromolar range, which is typical after primary selection from these libraries consisting of approximately 40 million different members each. Using these ligands in affinity chromatography, both target proteins were efficiently recovered from crude cell lysates with high selectivities. No loss of column capacity or selectivity was observed for repeated cycles of sample loading, washing and low pH elution. Interestingly, column sanitation could be performed using 0.5 M sodium hydroxide without significant loss of ligand performance. The results suggest that combinatorial approaches using robust protein domains as scaffolds can be a general tool in the process of designing purification strategies for biomolecules.

  • 339. Nord, K.
    et al.
    Nord, O.
    Uhlén, Mathias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Kelley, B.
    Ljungqvist, C.
    Nygren, Per-Åke
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Recombinant human factor VIII-specific affinity ligands selected from phage-displayed combinatorial libraries of protein A2001Inngår i: European Journal of Biochemistry, ISSN 0014-2956, E-ISSN 1432-1033, Vol. 268, nr 15, s. 4269-4277Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Factor VIII-specific affibodies were selected from phage displayed libraries constructed by combinatorial mutagenesis of an alpha helical bacterial receptor domain derived from staphylococcal protein A. Bead-immobilized recombinant human factor VIII (rVIII) (80 and 90 kDa chains) protein was used during competitive biopannings in the presence of free 80-kDa chain protein, resulting in the selection of several binders that showed dissociation constants (K-d) in the range 100-200 nm as determined by biosensor analyses. One variant (Z(rVIII:3), 90-kDa chain specific) was further characterized in small-scale affinity chromatography experiments, and showed efficient and selective recovery of biologically active rVIII from Chinese hamster ovary cell supernatant-derived feed stocks. The purity of the enriched rVIII was comparable with rVIII material purified by immunoaffinity chromatography using a 90-kDa chain-specific monoclonal antibody. Interestingly, epitope mapping showed that the monoclonal antibody and the affibody ligand competed for the same or at least overlapping epitopes on rVIII. In addition, the Z(rVIII:3) variant was produced by peptide synthesis with a C-terminal cysteine to enable directed coupling to solid supports. This 59-residue protein was analyzed by circular dichroism and showed a secondary structure content similar to that of the parental Z domain used as scaffold. In biosensor studies. the synthetic affibody was immobilized recruiting the C-terminal cysteine residue, and demonstrated to bind both recombinantly produced and plasma-derived factor VIII, From a secondary library, constructed by re-randomization of relevant positions identified after alignment of the first-generation variants, a panel of affinity-improved second-generation affibodies were selected of which one clone showed a dissociation constant (K-d) for rVIII of 5 nM. Several of these variants also showed higher apparent binding efficiencies towards rVIII when analyzed as immobilized ligands in biosensor experiments. Taken together, the results suggest that affibody ligands produced by bacterial or synthetic routes could be of interest as an alternative to monoclonal antibodies in purification processes or as diagnostic or monitoring tools.

  • 340. Nord, O.
    et al.
    Uhlén, Mathias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Nygren, Per-Åke
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Microbead display of proteins by cell-free expression of anchored DNA2003Inngår i: Journal of Biotechnology, ISSN 0168-1656, E-ISSN 1873-4863, Vol. 106, nr 1, s. 1-13Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Gene expression technologies where nucleic acid sequences remain physically linked to their corresponding gene products are important tools for selection and identification of rare variants in large protein libraries. Here, we describe a gene expression system, which combines the potential of bead-based suspension array technology (SAT) with gene expression and clonal identification. Using streptavidin-coated polystyrene micrometer-sized beads as solid supports for anchored PCR products, we have investigated conditions for cell-free expression and bioaffinity technology to provide clonal co-anchoring of corresponding gene products. Experiments showed that coupled transcription and translation of PCR product expression cassettes resulted in display of affinity-anchored proteins whose binding characteristics could be analyzed via direct and selective interaction with a fluorescently labeled target protein. Interestingly, experiments performed with differently biotinylated PCR products showed that the efficiency of display was dependent on the directionality of the expression cassette relative to the bead surface. In spiked systems, using small immunoglobulin binding proteins as models, we demonstrate efficient flow cytometric sorting of beads corresponding to the target interacting clones, verified by post-sorting analysis and clonal identification at DNA level. The use of this technology, including alternative for-mats, for different proteomics applications is discussed.

  • 341.
    Nygren, Per-Åke
    et al.
    KTH, Tidigare Institutioner, Bioteknologi.
    Uhlén, Mathias
    KTH, Tidigare Institutioner, Bioteknologi.
    Nord, Olof
    KTH, Tidigare Institutioner, Bioteknologi.
    In vitro selection and optional identification of polypeptides using solid support carriers1999Patent (Annet (populærvitenskap, debatt, mm))
    Abstract [en]

    The present invention relates to a nucleic acid molecule or a polypeptide that is selected from a method that includes cell-free expression of nucleic acid molecules immobilized on a sold support system. The present invention also relates to a molecular library that has a solid support system having a plurality of nucleic acid molecules immobilized thereon.

  • 342.
    Odeberg, Jacob
    et al.
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Ahmadian, Afshin
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Williams, C
    Pontén, F.
    Uhlén, Mathias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Lundeberg, Joakim
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Direct sequencing of the ZNF 189 gene promotor reveals artifact hot spot mutations.Manuskript (preprint) (Annet vitenskapelig)
  • 343.
    Odeberg, Jacob
    et al.
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Holmberg, K.
    Eriksson, P.
    Uhlén, Mathias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Molecular haplotyping by pyrosequencing (TM)2002Inngår i: BioTechniques, ISSN 0736-6205, E-ISSN 1940-9818, Vol. 33, nr 5, s. 1104-+Artikkel i tidsskrift (Fagfellevurdert)
  • 344.
    Odeberg, Jacob
    et al.
    KTH, Tidigare Institutioner (före 2005), Bioteknologi.
    Røsok, O
    Gudmundsson, G H
    Ahmadian, Afshin
    KTH, Tidigare Institutioner (före 2005), Bioteknologi.
    Roshani, L
    Williams, C
    Larsson, C
    Pontén, F
    Uhlén, Mathias
    KTH, Tidigare Institutioner (före 2005), Bioteknologi.
    Asheim, H C
    Lundeberg, Joakim
    KTH, Tidigare Institutioner (före 2005), Bioteknologi.
    Cloning and characterization of ZNF189, a novel human Krüppel-like zinc finger gene localized to chromosome 9q22-q31.1998Inngår i: Genomics, ISSN 0888-7543, E-ISSN 1089-8646, Vol. 50, nr 2, s. 213-21Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    A 3-kb-long cDNA encoding a Krüppel-like human zinc finger protein was isolated and mapped to chromosome 9q22-q31. The ZNF189 gene encodes a protein with 16 zinc fingers at its C-terminus and belongs to the Krüppel-associated box (KRAB)-containing group of zinc finger proteins. Four differently spliced cDNA transcripts, differing at the 5' coding region where a KRAB A repressor domain is encoded, were isolated. In addition, Northern blot analysis indicates the presence of two additional unidentified splice variants. Comparison of cDNA and genomic sequences shows that the ZNF189 gene spans approximately 11 kb and is organized into at least four exons, the large 3'-end exon coding for the complete zinc finger domain and the 3' untranslated region. ZNF189 is expressed in all tissues and cell types currently investigated, at varying levels, but with a tissue- or cell-type-restricted expression pattern for the different splice variants. ZNF189 is conserved in the genome of several mammalian species. Direct sequencing of the ZNF189 gene in microdissected tumor biopsies of sporadic basal cell carcinoma and squamous cell carcinoma reveals no mutations in the coding sequence or at exon/intron boundaries.

  • 345. O'Hurley, Gillian
    et al.
    Busch, Christer
    Fagerberg, Linn
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Hallström, Björn M.
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Stadler, Charlotte
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Tolf, Anna
    Lundberg, Emma
    Schwenk, Jochen M.
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Jirstrom, Karin
    Bjartell, Anders
    Gallagher, William M.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Ponten, Fredrik
    Analysis of the Human Prostate-Specific Proteome Defined by Transcriptomics and Antibody-Based Profiling Identifies TMEM79 and ACOXL as Two Putative, Diagnostic Markers in Prostate Cancer2015Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, nr 8, artikkel-id e0133449Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    To better understand prostate function and disease, it is important to define and explore the molecular constituents that signify the prostate gland. The aim of this study was to define the prostate specific transcriptome and proteome, in comparison to 26 other human tissues. Deep sequencing of mRNA (RNA-seq) and immunohistochemistry-based protein profiling were combined to identify prostate specific gene expression patterns and to explore tissue biomarkers for potential clinical use in prostate cancer diagnostics. We identified 203 genes with elevated expression in the prostate, 22 of which showed more than five-fold higher expression levels compared to all other tissue types. In addition to previously well-known proteins we identified two poorly characterized proteins, TMEM79 and ACOXL, with potential to differentiate between benign and cancerous prostatic glands in tissue biopsies. In conclusion, we have applied a genome-wide analysis to identify the prostate specific proteome using transcriptomics and antibody-based protein profiling to identify genes with elevated expression in the prostate. Our data provides a starting point for further functional studies to explore the molecular repertoire of normal and diseased prostate including potential prostate cancer markers such as TMEM79 and ACOXL.

  • 346. O'Leary, Patrick C.
    et al.
    Penny, Sarah A.
    Dolan, Roisin T.
    Kelly, Catherine M.
    Madden, Stephen F.
    Rexhepaj, Elton
    Brennan, Donal J.
    McCann, Amanda H.
    Ponten, Fredrik
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik.
    Zagozdzon, Radoslaw
    Duffy, Michael J.
    Kell, Malcolm R.
    Jirstrom, Karin
    Gallagher, William M.
    Systematic antibody generation and validation via tissue microarray technology leading to identification of a novel protein prognostic panel in breast cancer2013Inngår i: BMC Cancer, ISSN 1471-2407, E-ISSN 1471-2407, Vol. 13Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: Although omic-based discovery approaches can provide powerful tools for biomarker identification, several reservations have been raised regarding the clinical applicability of gene expression studies, such as their prohibitive cost. However, the limited availability of antibodies is a key barrier to the development of a lower cost alternative, namely a discrete collection of immunohistochemistry (IHC)-based biomarkers. The aim of this study was to use a systematic approach to generate and screen affinity-purified, mono-specific antibodies targeting progression-related biomarkers, with a view towards developing a clinically applicable IHC-based prognostic biomarker panel for breast cancer. Methods: We examined both in-house and publicly available breast cancer DNA microarray datasets relating to invasion and metastasis, thus identifying a cohort of candidate progression-associated biomarkers. Of these, 18 antibodies were released for extended analysis. Validated antibodies were screened against a tissue microarray (TMA) constructed from a cohort of consecutive breast cancer cases (n = 512) to test the immunohistochemical surrogate signature. Results: Antibody screening revealed 3 candidate prognostic markers: the cell cycle regulator, Anillin (ANLN); the mitogen-activated protein kinase, PDZ-Binding Kinase (PBK); and the estrogen response gene, PDZ-Domain Containing 1 (PDZK1). Increased expression of ANLN and PBK was associated with poor prognosis, whilst increased expression of PDZK1 was associated with good prognosis. A 3-marker signature comprised of high PBK, high ANLN and low PDZK1 expression was associated with decreased recurrence-free survival (p < 0.001) and breast cancer-specific survival (BCSS) (p < 0.001). This novel signature was associated with high tumour grade (p < 0.001), positive nodal status (p = 0.029), ER-negativity (p = 0.006), Her2-positivity (p = 0.036) and high Ki67 status (p < 0.001). However, multivariate Cox regression demonstrated that the signature was not a significant predictor of BCSS (HR = 6.38; 95% CI = 0.79-51.26, p = 0.082). Conclusions: We have developed a comprehensive biomarker pathway that extends from discovery through to validation on a TMA platform. This proof-of-concept study has resulted in the identification of a novel 3-protein prognostic panel. Additional biochemical markers, interrogated using this high-throughput platform, may further augment the prognostic accuracy of this panel to a point that may allow implementation into routine clinical practice.

  • 347. O'Leary, Patrick C.
    et al.
    Terrile, Marta
    Bajor, Malgorzata
    Gaj, Pawel
    Hennessy, Bryan T.
    Mills, Gordon B.
    Zagozdzon, Agnieszka
    O'Connor, Darran P.
    Brennan, Donal J.
    Connor, Kate
    Li, Jane
    Gonzalez-Angulo, Ana Maria
    Sun, Han-Dong
    Pu, Jian-Xin
    Pontén, Fredrik
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
    Jirstrom, Karin
    Nowis, Dominika A.
    Crown, John P.
    Zagozdzon, Radoslaw
    Gallagher, William M.
    Peroxiredoxin-1 protects estrogen receptor alpha from oxidative stress-induced suppression and is a protein biomarker of favorable prognosis in breast cancer2014Inngår i: Breast Cancer Research, ISSN 1465-5411, E-ISSN 1465-542X, Vol. 16, nr 4, s. R79-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Introduction: Peroxiredoxin-1 (PRDX1) is a multifunctional protein, acting as a hydrogen peroxide (H2O2) scavenger, molecular chaperone and immune modulator. Although differential PRDX1 expression has been described in many tumors, the potential role of PRDX1 in breast cancer remains highly ambiguous. Using a comprehensive antibody-based proteomics approach, we interrogated PRDX1 protein as a putative biomarker in estrogen receptor (ER)-positive breast cancer. Methods: An anti-PRDX1 antibody was validated in breast cancer cell lines using immunoblotting, immunohistochemistry and reverse phase protein array (RPPA) technology. PRDX1 protein expression was evaluated in two independent breast cancer cohorts, represented on a screening RPPA (n = 712) and a validation tissue microarray (n = 498). In vitro assays were performed exploring the functional contribution of PRDX1, with oxidative stress conditions mimicked via treatment with H2O2, peroxynitrite, or adenanthin, a PRDX1/2 inhibitor. Results: In ER-positive cases, high PRDX1 protein expression is a biomarker of improved prognosis across both cohorts. In the validation cohort, high PRDX1 expression was an independent predictor of improved relapse-free survival (hazard ratio (HR) = 0.62, 95% confidence interval (CI) = 0.40 to 0.96, P = 0.032), breast cancer-specific survival (HR = 0.44, 95% CI = 0.24 to 0.79, P = 0.006) and overall survival (HR = 0.61, 95% CI = 0.44 to 0.85, P = 0.004). RPPA screening of cancer signaling proteins showed that ER alpha protein was upregulated in PRDX1 high tumors. Exogenous H2O2 treatment decreased ER alpha protein levels in ER-positive cells. PRDX1 knockdown further sensitized cells to H2O2- and peroxynitrite-mediated effects, whilst PRDX1 overexpression protected against this response. Inhibition of PRDX1/2 antioxidant activity with adenanthin dramatically reduced ER alpha levels in breast cancer cells. Conclusions: PRDX1 is shown to be an independent predictor of improved outcomes in ER-positive breast cancer. Through its antioxidant function, PRDX1 may prevent oxidative stress-mediated ER alpha loss, thereby potentially contributing to maintenance of an ER-positive phenotype in mammary tumors. These results for the first time imply a close connection between biological activity of PRDX1 and regulation of estrogen-mediated signaling in breast cancer.

  • 348. Olofsson, S. -E
    et al.
    Nodin, B.
    Gaber, A.
    Eberhard, J.
    Uhlén, Mathias
    KTH, Centra, Science for Life Laboratory, SciLifeLab. KTH, Skolan för bioteknologi (BIO).
    Jirström, K.
    Jerkeman, M.
    Low RBM3 protein expression correlates with clinical stage, prognostic classification and increased risk of treatment failure in testicular non-seminomatous germ cell cancer2015Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, nr 3, artikkel-id e0121300Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: Expression of the RNA-binding motif protein 3 (RBM3) has been shown to correlate with favourable clinicopathological parameters and prognosis in several cancer diseases. The aim of this study was to examine the expression and prognostic ability of RBM3 in patients with testicular non-seminomatous germ cell tumours (NSGCT). Patients and Methods: Immunohistochemical RBM3 expression was analysed in tissue microarrays with tumours from 206 patients. Chi-square test was applied to analyze associations between RBM3 expression and clinicopathological parameters. Kaplan-Meier analysis was used to assess the impact of RBM3 expression on cancer-specific survival (CSS) and failure-free survival (FFS). Cox regression proportional hazards models were used to estimate the relative risk for failure. Results: In the entire cohort, there was a significant association between clinical stage (p=0.044) and RBM3 expression. Weak RBM3 expression correlated with a significantly reduced FFS [79.3% versus 90.4%(p=0.019)] and CSS [87.5% versus 97.3% (p=0.047)]. For patients with metastatic disease (n = 88), significant associations were found between RBM3 expression and IGCCC group (p=0.007). The FFS was significantly inferior for patients with low tumour-specific RBM3 expression [59.3% versus 79.0% (p=0.013)], and this association remained significant in a multivariable model for patients with metastatic disease (HR=3.67; 95% CI 1.14, 11.89). Conclusion: Low RBM3 expression is an independent predictor of treatment failure in metastatic NSGCT, in relation to the prognostic factors included in the International Germ Cell Consensus Classification (IGCCC). These findings suggest that RBM3 may be a potential biomarker for treatment stratification in patients with metastatic non-seminomatous germ cell tumours, and therefore merit further validation.

  • 349.
    O'Meara, Deirdre
    et al.
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Nilsson, Peter
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Nygren, Per-Åke
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Uhlen, Mathias
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Lundeberg, Joakim
    KTH, Tidigare Institutioner                               , Bioteknologi.
    Capture of single-stranded DNA assisted by oligonucleotide modules1998Inngår i: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 255, nr 2, s. 195-203Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Real-time biospecific interaction analysis was employed to monitor direct capture of a hepatitis C virus (HCV) derived polymerase chain reaction (PCR) product by nucleic acid hybridization. Different formats for hybridization were used to study the interaction between a single-stranded HCV PCR product and capture oligonucleotides immobilized on a sensor chip via streptavidin-biotin chemistry. By employing a prehybridization step in solution with nonbiotin oligonucleotides complementary to the single-stranded target and adjacent to the immobilized probe, a significant capture was achieved in comparison to the low capture efficiency obtained using single immobilized probes (9-36 mer). High capture efficiencies were also observed when shorter immobilized probes were used in combination with strings of adjacently positioned prehybridized probes (i.e., modules). Interestingly, the introduction of single nucleotide gaps between prehybridized and/or immobilized probes dramatically reduced the capture efficiency. These results suggest that flexible systems for capture could be designed from libraries of short oligonucleotides (9 mers) used in module fashion, taking advantage of stacking interactions between the oligonucleotides. The potential applications of such oligonucleotide-assisted capture systems are discussed.

  • 350. Orchard, S.
    et al.
    Heck, A.
    Uhlén, Mathias
    KTH, Skolan för bioteknologi (BIO), Proteomik (stängd 20130101).
    Ping, P.
    Publication Committee Meeting HUPO 5th Annual World Congress Long Beach, CA, USA 30 October 20062007Inngår i: Proteomics, ISSN 1615-9853, E-ISSN 1615-9861, Vol. 7, nr 7, s. 1009-1011Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    This meeting brought together delegates from industry, academia and the publishing houses to facilitate discussions on the level of support from the journals for the use of standardised data formats and their interest in the creation of a network of proteomics repositories collaborating on a coordinated data curation effort. Discussions centred on how best to structure interactions between journals, databases and researchers to improve accessibility to data, and facilitate comparisons between datasets.

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