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  • 1. Bjork, L.
    et al.
    Ait Blal, C.
    Alm, Tove L.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Bäckström, Anna
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Gnann, C.
    Hjelmare, Martin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Schutten, Rutger
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Stadler, Charlotte
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Application specific antibody validation. The Human Protein Atlas validation scheme and how to confirm subcellular protein localization.2016In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 27Article in journal (Refereed)
  • 2.
    Danielsson, Frida
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Akesson, L.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Profiling changes in response to hypoxia in a four-step cell line model for malignant transformation.2016In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 27Article in journal (Refereed)
  • 3.
    Danielsson, Frida
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Huss, Mikael
    Rexhepaj, Elton
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    O'Hurley, Gillian
    Klevebring, Daniel
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Pontén, Fredrik
    Gad, Annica K. B.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Majority of differentially expressed genes are down-regulated during malignant transformation in a four-stage model2013In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 110, no 17, p. 6853-6858Article in journal (Refereed)
    Abstract [en]

    The transformation of normal cells to malignant, metastatic tumor cells is a multistep process caused by the sequential acquirement of genetic changes. To identify these changes, we compared the transcriptomes and levels and distribution of proteins in a four-stage cell model of isogenically matched normal, immortalized, transformed, and metastatic human cells, using deep transcriptome sequencing and immunofluorescence microscopy. The data show that similar to 6% (n = 1,357) of the human protein-coding genes are differentially expressed across the stages in the model. Interestingly, the majority of these genes are down-regulated, linking malignant transformation to dedifferentiation. The up-regulated genes are mainly components that control cellular proliferation, whereas the down-regulated genes consist of proteins exposed on or secreted from the cell surface. As many of the identified gene products control basic cellular functions that are defective in cancers, the data provide candidates for follow-up studies to investigate their functional roles in tumor formation. When we further compared the expression levels of four of the identified proteins in clinical cancer cohorts, similar differences were observed between benign and cancer cells, as in the cell model. This shows that this comprehensive demonstration of the molecular changes underlying malignant transformation is a relevant model to study the process of tumor formation.

  • 4.
    Danielsson, Frida
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Åkesson, L.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Mahdessian, Diana
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Sullivan, Devin P.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Thul, Peter
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Wiking, Mikaela
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Björk, L.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Schutten, Rutger
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Ait Blal, Carl
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Hjelmare, Martin
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Gnann, Christian
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    An image-based view of the microtubule proteome2016In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 27Article in journal (Refereed)
  • 5.
    Danielsson, Frida
    et al.
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Wiking, Mikaela
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mahdessian, Diana
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Ait Blal, Hammou
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hjelmare, Martin
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Stadler, Charlotte
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    RNA Deep Sequencing as a Tool for Selection of Cell Lines for Systematic Subcellular Localization of All Human Proteins2013In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 12, no 1, p. 231-239Article in journal (Refereed)
    Abstract [en]

    One of the major challenges of a chromosome-centric proteome project is to explore in a systematic manner the potential proteins identified from the chromosomal genome sequence, but not yet characterized on a protein level. Here, we describe the use of RNA deep sequencing to screen human cell lines for RNA profiles and to use this information to select cell lines suitable for characterization of the corresponding gene product. In this manner, the subcellular localization of proteins can be analyzed systematically using antibody-based confocal microscopy. We demonstrate the usefulness of selecting cell lines with high expression levels of RNA transcripts to increase the likelihood of high quality immunofluorescence staining and subsequent successful subcellular localization of the corresponding protein. The results show a path to combine transcriptomics with affinity proteomics to characterize the proteins in a gene- or chromosome-centric manner.

  • 6.
    Fagerberg, Linn
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hallström, Björn M.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Oksvold, Per
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Kampf, C.
    Djureinovic, D.
    Odeberg, Jacob
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Habuka, Masato
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Tahmasebpoor, S.
    Danielsson, A.
    Edlund, K.
    Asplund, A.
    Sjöstedt, E.
    Lundberg, E.
    Szigyarto, Cristina Al-Khalili
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Ottosson Takanen, J.
    Berling, H.
    Tegel, Hanna
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Mulder, J.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lindskog, C.
    Danielsson, Frida
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mardinoglu, A.
    Sivertsson, Åsa
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Von Feilitzen, Kalle
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Forsberg, Mattias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Zwahlen, Martin
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Olsson, I.
    Navani, S.
    Huss, Mikael
    Nielsen, Jens
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Pontén, F.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Analysis of the human tissue-specific expression by genome-wide integration of transcriptomics and antibody-based proteomics2014In: Molecular & Cellular Proteomics, ISSN 1535-9476, E-ISSN 1535-9484, Vol. 13, no 2, p. 397-406Article in journal (Refereed)
    Abstract [en]

    Global classification of the human proteins with regards to spatial expression patterns across organs and tissues is important for studies of human biology and disease. Here, we used a quantitative transcriptomics analysis (RNA-Seq) to classify the tissue-specific expression of genes across a representative set of all major human organs and tissues and combined this analysis with antibody- based profiling of the same tissues. To present the data, we launch a new version of the Human Protein Atlas that integrates RNA and protein expression data corresponding to 80% of the human protein-coding genes with access to the primary data for both the RNA and the protein analysis on an individual gene level. We present a classification of all human protein-coding genes with regards to tissue-specificity and spatial expression pattern. The integrative human expression map can be used as a starting point to explore the molecular constituents of the human body.

  • 7.
    Fagerberg, Linn
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Oksvold, Per
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Älgenäs, C.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Pontén, F.
    Sivertsson, Åsa
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Odeberg, Jacob
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Klevebring, Daniel
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Kampf, C.
    Asplund, A.
    Sjöstedt, E.
    Al-Khalili Szigyarto, C.
    Edqvist, P. -H
    Olsson, I.
    Rydberg, U.
    Hudson, P.
    Ottosson Takanen, J.
    Berling, H.
    Björling, Lisa
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Tegel, Hanna
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Rockberg, J.
    Nilsson, Peter
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Navani, S.
    Jirström, K.
    Mulder, J.
    Schwenk, Jochen M.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Zwahlen, Martin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hober, Sophia
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Forsberg, Mattias
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Von Feilitzen, Kalle
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Contribution of antibody-based protein profiling to the human chromosome-centric proteome project (C-HPP)2013In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 12, no 6, p. 2439-2448Article in journal (Refereed)
    Abstract [en]

    A gene-centric Human Proteome Project has been proposed to characterize the human protein-coding genes in a chromosome-centered manner to understand human biology and disease. Here, we report on the protein evidence for all genes predicted from the genome sequence based on manual annotation from literature (UniProt), antibody-based profiling in cells, tissues and organs and analysis of the transcript profiles using next generation sequencing in human cell lines of different origins. We estimate that there is good evidence for protein existence for 69% (n = 13985) of the human protein-coding genes, while 23% have only evidence on the RNA level and 7% still lack experimental evidence. Analysis of the expression patterns shows few tissue-specific proteins and approximately half of the genes expressed in all the analyzed cells. The status for each gene with regards to protein evidence is visualized in a chromosome-centric manner as part of a new version of the Human Protein Atlas (www.proteinatlas.org).

  • 8.
    Fagerberg, Linn
    et al.
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Sandler, Charlotte
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Skogs, Marie
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hjelmare, Martin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Jonasson, Kalle
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Wiking, Mikaela
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Åbergh, Annica
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundberg, Emma
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mapping the subcellular protein distribution in three human cell lines2011In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 10, no 8, p. 3766-3777Article in journal (Refereed)
    Abstract [en]

    The subcellular locations of proteins are closely related to their function and constitute an essential aspect for understanding the complex machinery of living cells. A systematic effort has been initiated to map the protein distribution in three functionally different cell lines with the aim to provide a subcellular localization index for at least one representative protein from all human protein-encoding genes. Here, we present the results of over 4,000 proteins mapped to 16 subcellular compartments. The results indicate a ubiquitous protein expression with a majority of the proteins found in all three cell lines and a large portion localized to two or more compartments. The inter-relationships between the subcellular compartments are visualized in a protein-compartment network based on all detected proteins. Hierarchical clustering was performed to determine how closely related the organelles are in terms of protein constituents and compare the proteins detected in each cell type. Our results show distinct organelle proteomes, well conserved across the cell types, and demonstrate that biochemically similar organelles are grouped together.

  • 9.
    Grimm, Sebastian
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics.
    Yu, Feifan
    KTH, School of Biotechnology (BIO), Proteomics.
    Shibasaki, Seiji
    Vernet, Erik
    KTH, School of Biotechnology (BIO), Proteomics.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics.
    Nygren, Per-Åke
    KTH, School of Biotechnology (BIO), Molecular Biotechnology.
    Gräslund, Torbjörn
    KTH, School of Biotechnology (BIO), Molecular Biotechnology.
    Selection and characterisation of affibody molecules inhibiting the interaction between Ras and Raf in vitro2010In: NEW BIOTECHNOL, ISSN 1871-6784, Vol. 27, no 6, p. 766-773Article in journal (Refereed)
    Abstract [en]

    Development of molecules with the ability to selectively inhibit particular protein-protein interactions is important in providing tools for understanding cell biology In this work, we describe efforts to select small Ras- and Raf-specific three-helix bundle affibody binding proteins capable of inhibiting the interaction between H-Ras and Raf-1, from a combinatorial library displayed on bacteriophage Target-specific variants with typically high nanomolar or low micromolar affinities (K-D) could be selected successfully against both proteins, as shown by dot blot, ELISA and real-time biospecific interaction analyses Affibody molecule variants selected against H-Ras were shown to bind epitopes overlapping each other at a site that differed from that at which H-Ras interacts with Raf-1 In contrast, an affibody molecule isolated during selection against Raf-1 was shown to effectively inhibit the interaction between H-Ras and Raf-1 in a dose-dependent manner Possible intracellular applications of the selected affibody molecules are discussed

  • 10. Jakobsen, Lis
    et al.
    Vanselow, Katja
    Skogs, Marie
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Toyoda, Yusuke
    Lundberg, Emma
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Poser, Ina
    Falkenby, Lasse G.
    Bennetzen, Martin
    Westendorf, Jens
    Nigg, Erich A.
    Uhlen, Mathias
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Hyman, Anthony A.
    Andersen, Jens S.
    Novel asymmetrically localizing components of human centrosomes identified by complementary proteomics methods2011In: EMBO Journal, ISSN 0261-4189, E-ISSN 1460-2075, Vol. 30, no 8, p. 1520-1535Article in journal (Refereed)
    Abstract [en]

    Centrosomes in animal cells are dynamic organelles with a proteinaceous matrix of pericentriolar material assembled around a pair of centrioles. They organize the microtubule cytoskeleton and the mitotic spindle apparatus. Mature centrioles are essential for biogenesis of primary cilia that mediate key signalling events. Despite recent advances, the molecular basis for the plethora of processes coordinated by centrosomes is not fully understood. We have combined protein identification and localization, using PCP-SILAC mass spectrometry, BAC transgeneOmics, and antibodies to define the constituents of human centrosomes. From a background of non-specific proteins, we distinguished 126 known and 40 candidate centrosomal proteins, of which 22 were confirmed as novel components. An antibody screen covering 4000 genes revealed an additional 113 candidates. We illustrate the power of our methods by identifying a novel set of five proteins preferentially associated with mother or daughter centrioles, comprising genes implicated in cell polarity. Pulsed labelling demonstrates a remarkable variation in the stability of centrosomal protein complexes. These spatiotemporal proteomics data provide leads to the further functional characterization of centrosomal proteins.

  • 11.
    Mahdessian, Diana
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Wiking, Mikaela
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Åkesson, Lars
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Danielsson, Frida
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Ait Blal, Carl
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sullivan, Devin P.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Thul, Peter
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Gnann, Christian
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Bäckström, Anna
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Fall, Jenny
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Schutten, Rutger
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Björk, Lars
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hjelmare, Martin
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Stadler, Charlotte
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Profiling the human cytoplasmic proteome.2016In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 27Article in journal (Refereed)
  • 12.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. Science for Life Laboratory / Royal Institute of Technology.
    Antibody-based subcellular localization of the human proteome2016Licentiate thesis, comprehensive summary (Other academic)
    Abstract [en]

    This thesis describes the use of antibodies and immunofluorescence for subcellular localization of proteins. The key objective is the creation of an open-source atlas with information on the subcellular location of every human protein. Knowledge of the spatial distribution and the precise location of a protein within a cell is important for its functional characterization, and describing the human proteome in terms of compartment proteomes is important to decipher cellular organization and function.

     

    Immunofluorescence and confocal microscopy of cultured cells were used for high-resolution detection of proteins on a high-throughput scale. Critical to immunofluorescence results are sample preparation and specific antibodies. Antibody staining of cells requires fixation and permeabilization, both of which can result in loss or redistribution of proteins and masking of epitopes. A high-throughput approach demands a standardized protocol suitable for the majority of proteins across cellular compartments. Paper I presents an evaluation of sample preparation techniques from which such a single fixation and permeabilization protocol was optimized. Paper II describes the results from applying this protocol to 4000 human proteins in three cell lines of different origin.

     

    Paper III presents a strategy for application-specific antibody validation. Antibodies are the key reagents in immunofluorescence, but all antibodies have potential for off-target binding and should be validated thoroughly. Antibody performance varies across sample types and applications due to the competition present and the effect of the sample preparation on antigen accessibility. In this paper application-specific validation for immunofluorescence was conducted using colocalization with fluorescently tagged protein in transgenic cell lines. 

  • 13.
    Skogs, Marie
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab. Royal Inst Technol, Sci Life Lab, Stockholm, Sweden..
    Lundberg, Emma
    KTH, Centres, Science for Life Laboratory, SciLifeLab. Royal Inst Technol, Sci Life Lab, Stockholm, Sweden..
    Proteins that assemble into Rods & Rings - subcellular protein complexes with unknown functions.2015In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 26Article in journal (Other academic)
  • 14.
    Skogs, Marie
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Stadler, Charlotte
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Schutten, Rutger
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hjelmare, Martin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Gnann, Christian
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Björk, Lars
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Poser, Ina
    Hyman, Anthony
    Uhlén, Mathias
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundberg, Emma
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Antibody Validation in Bioimaging Applications Based on Endogenous Expression of Tagged Proteins2017In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 16, no 1, p. 147-155Article in journal (Refereed)
    Abstract [en]

    Antibodies are indispensible research tools, yet the scientific community has not adopted standardized procedures to validate their specificity. Here we present a strategy to systematically validate antibodies for immunofluorescence (IF) applications using gene tagging. We have assessed the on- and off-target binding capabilities of 197 antibodies using 108 cell lines expressing EGFP-tagged target proteins at endogenous levels. Furthermore, we assessed batch-to-batch effects for 35 target proteins, showing that both the on- and off-target binding patterns vary significantly between antibody batches and that the proposed strategy serves as a reliable procedure for ensuring reproducibility upon production of new antibody batches. In summary, we present a systematic scheme for antibody validation in IF applications using endogenous expression of tagged proteins. This is an important step toward a reproducible approach for context- and application-specific antibody validation and improved reliability of antibody-based experiments and research data.

  • 15.
    Skogs, Marie
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. Science for Life Laboratory / Royal Institute of Technology.
    Stadler, Charlotte
    Schutten, Rutger
    Hjelmare, Martin
    Gnann, Christian
    Poser, Ina
    Hyman, Anthony
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Lundberg, Emma
    An antibody validation scheme for immunofluorescence using gene taggingManuscript (preprint) (Other academic)
  • 16.
    Stadler, Charlotte
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics.
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics.
    A single fixation protocol for proteome-wide immunofluorescence localization studies2010In: Journal of Proteomics, ISSN 1874-3919, Vol. 73, no 6, p. 1067-1078Article in journal (Refereed)
    Abstract [en]

    Immunofluorescence microscopy is a valuable tool for analyzing protein expression and localization at a subcellular level thus providing information regarding protein function, interaction partners and its role in cellular processes. When performing sample fixation, parameters such as difference in accessibility of proteins present in various cellular compartments as well as the chemical composition of the protein to be studied, needs to be taken into account. However, in systematic and proteome-wide efforts, a need exists for standard fixation protocol(s) that works well for the majority of all proteins independent of subcellular localization. Here, we report on a study with the goal to find a standardized protocol based on the analysis of 18 human proteins localized in 11 different organelles and subcellular structures. Six fixation protocols were tested based on either dehydration by alcohols (methanol, ethanol or iso-propanol) or cross-linking by paraformaldehyde followed by detergent permeabilization (Triton X-100 or saponin) in three human cell lines. Our results show that cross-linking is essential for proteome-wide localization studies and that cross-linking using paraformaldehyde followed by Triton X-100 permeabilization successfully can be used as a single fixation protocol for systematic studies.

  • 17.
    Thul, Peter J.
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Åkesson, Lovisa
    KTH, School of Biotechnology (BIO). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Wiking, Mikaela
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mahdessian, Diana
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Geladaki, A.
    Ait Blal, Hammou
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Alm, Tove L.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Asplund, A.
    Björk, Lars
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Breckels, L. M.
    Bäckström, Anna
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Danielsson, Frida
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Fagerberg, Linn
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Fall, Jenny
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Gatto, L.
    Gnann, Christian
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Protein Technology.
    Hjelmare, Martin
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Johansson, Fredric
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lee, Sunjae
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lindskog, C.
    Mulder, J.
    Mulvey, C. M.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Oksvold, Per
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Rockberg, Johan
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Schutten, Rutger
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sivertsson, Åsa
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sjöstedt, E.
    Skogs, Marie
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Stadler, Charlotte
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sullivan, Devin P.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Tegel, Hanna
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Winsnes, Casper F.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Zhang, Cheng
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Zwahlen, Martin
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mardinoglu, Adil
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Pontén, F.
    von Feilitzen, Kalle
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lilley, K. S.
    Uhlén, Mathias
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Lundberg, Emma
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    A subcellular map of the human proteome2017In: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 356, no 6340, article id 820Article in journal (Refereed)
    Abstract [en]

    Resolving the spatial distribution of the human proteome at a subcellular level can greatly increase our understanding of human biology and disease. Here we present a comprehensive image-based map of subcellular protein distribution, the Cell Atlas, built by integrating transcriptomics and antibody-based immunofluorescence microscopy with validation by mass spectrometry. Mapping the in situ localization of 12,003 human proteins at a single-cell level to 30 subcellular structures enabled the definition of the proteomes of 13 major organelles. Exploration of the proteomes revealed single-cell variations in abundance or spatial distribution and localization of about half of the proteins to multiple compartments. This subcellular map can be used to refine existing protein-protein interaction networks and provides an important resource to deconvolute the highly complex architecture of the human cell.

  • 18.
    Wiking, Mikaela
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Skogs, Marie
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hjelmare, Martin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Blal, Hammou Ait
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Stadler, Charlotte
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mahdessian, Diana
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Danielsson, Frida
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Schutten, Rutger
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Åbergh, Annica
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Uhlén, Mathias
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundberg, Emma
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    The Subcellular Protein Atlas2014In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 25, article id P1603Article in journal (Other academic)
1 - 18 of 18
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