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  • 1. Bashiruddin, J. B.
    et al.
    de Santis, P.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Ball, H.
    Regalla, J.
    Detection of Mycoplasma mycoides subspecies mycoides SC in bovine lung and lymph node tissues by culture, sandwich ELISA and polymerase chain reaction systems2005In: Research in Veterinary Science, ISSN 0034-5288, E-ISSN 1532-2661, Vol. 78, no 3, p. 199-205Article in journal (Refereed)
    Abstract [en]

    Cattle from Northern Portugal, many with pulmonary lesions typical of contagious bovine pleuropneumonia, were investigated for the presence of Mycoplasma mycoides subspecies mycoides small colony (MmmSC), which is the causative agent of CBPP, with several detection tests. sandwich ELISA that included a Culture enrichment stage, and 2 different PCR diagnostic systems were used to detect MmmSC in lung and mediastinal lymph node tissues from these animals. The comparison of typical CBPP pathology with the results of detection revealed that no single one of these methods provided a perfect match to the pathological data. Best performing tests were the PCR with laser induced fluorescence and PCR with pleuroTRAP kit (Chemicon, Australia), which are diagnostic systems based on amplification of genomic MmmSC DNA Followed by sensitive detection of the amplified products. These were followed by the broth-enriched sandwich ELISA, Which uses a monoclonal antibody specific to the M. mycoides cluster, to capture the antigen.

  • 2.
    Berglund, Lisa
    et al.
    KTH, School of Biotechnology (BIO).
    Björling, Erik
    KTH, School of Biotechnology (BIO).
    Gry, Marcus
    KTH, School of Biotechnology (BIO).
    Asplund, Anna
    Uppsala Univ, Rudbeck laboratory.
    Al-Khalili Szigyarto, Cristina
    KTH, School of Biotechnology (BIO).
    Persson, Anja
    KTH, School of Biotechnology (BIO).
    Ottoson, Jenny
    KTH, School of Biotechnology (BIO).
    Wernérus, Henrik
    KTH, School of Biotechnology (BIO).
    Nilsson, Peter
    KTH, School of Biotechnology (BIO).
    Sivertsson, Åsa
    KTH, School of Biotechnology (BIO).
    Wester, Kenneth
    Uppsala Univ, Rudbeck laboratory.
    Kampf, Caroline
    Uppsala Univ, Rudbeck laboratory.
    Hober, Sophia
    KTH, School of Biotechnology (BIO).
    Pontén, Fredrik
    Uppsala Univ, Rudbeck laboratory.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO).
    Generation of validated antibodies towards the human proteomeArticle in journal (Other academic)
    Abstract [en]

    Here we show the results from a large effort to generate antibodies towards the human proteome. A high-throughput strategy was developed based on cloning and expression of antigens as recombitant protein epitope signature tags (PrESTs) Affinity purified polyclonal antibodies were generated, followed by validation by protein microarrays, Western blotting and microarray-based immunohistochemistry. PrESTs were selected based on sequence uniqueness relative the proteome and a bioinformatics analysis showed that unique antigens can be found for at least 85% of the proteome using this general strategy. The success rate from antigen selection to validated antibodies was 31%, and from protein to antibody 55%. Interestingly, membrane-bound and soluble proteins performed equally and PrEST lengths between 75 and 125 amino acids were found to give the highest yield of validated antibodies. Multiple antigens were selected for many genes and the results suggest that specific antibodies can be systematically generated to most human proteibs.

  • 3.
    Berglund, Lisa
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Björling, Erik
    KTH, School of Biotechnology (BIO), Proteomics.
    Oksvold, Per
    KTH, School of Biotechnology (BIO), Proteomics.
    Fagerberg, Linn
    KTH, School of Biotechnology (BIO), Proteomics.
    Al-Khalili Szigyarto, Cristina
    KTH, School of Biotechnology (BIO), Proteomics.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Ottosson, Jenny
    KTH, School of Biotechnology (BIO), Proteomics.
    Wernérus, Henrik
    KTH, School of Biotechnology (BIO), Proteomics.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics.
    Sivertsson, Åsa
    KTH, School of Biotechnology (BIO), Proteomics.
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Proteomics.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    et al.,
    A genecentric human protein atlas for expression profiles based on antibodies2008In: Molecular & Cellular Proteomics, ISSN 1535-9476, Vol. 7, no 10, p. 2019-2027Article in journal (Refereed)
    Abstract [en]

    An attractive path forward in proteomics is to experimentally annotate the human protein complement of the genome in a genecentric manner. Using antibodies, it might be possible to design protein-specific probes for a representative protein from every protein-coding gene and to subsequently use the antibodies for systematical analysis of cellular distribution and subcellular localization of proteins in normal and disease tissues. A new version (4.0) of the Human Protein Atlas has been developed in a genecentric manner with the inclusion of all human genes and splice variants predicted from genome efforts together with a visualization of each protein with characteristics such as predicted membrane regions, signal peptide, and protein domains and new plots showing the uniqueness (sequence similarity) of every fraction of each protein toward all other human proteins. The new version is based on tissue profiles generated from 6120 antibodies with more than five million immunohistochemistry-based images covering 5067 human genes, corresponding to similar to 25% of the human genome. Version 4.0 includes a putative list of members in various protein classes, both functional classes, such as kinases, transcription factors, G-protein-coupled receptors, etc., and project-related classes, such as candidate genes for cancer or cardiovascular diseases. The exact antigen sequence for the internally generated antibodies has also been released together with a visualization of the application-specific validation performed for each antibody, including a protein array assay, Western blot analysis, immunohistochemistry, and, for a large fraction, immunofluorescence-based confocal microscopy. New search functionalities have been added to allow complex queries regarding protein expression profiles, protein classes, and chromosome location. The new version of the protein atlas thus is a resource for many areas of biomedical research, including protein science and biomarker discovery.

  • 4.
    Berglund, Lisa
    et al.
    KTH, School of Biotechnology (BIO).
    Persson, Anja
    KTH, School of Biotechnology (BIO).
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO).
    Primer design for high-throughput PCR cloningArticle in journal (Other academic)
  • 5.
    Beven, Laure
    et al.
    Univ Bordeaux, Villenave Dornon, France ; INRA Villenave Dornon, France .
    Charenton, Claire
    Univ Bordeaux, Villenave Dornon, France ; INRA Villenave Dornon, France .
    Dautant, Alain
    Univ Bordeaux, Bordeaux, France ; IBMC, CNRS, Bordeaux, France.
    Bouyssou, Guillaume
    Univ Bordeaux, Villenave Dornon, France ; INRA Villenave Dornon, France .
    Labroussaa, Fabien
    Univ Bordeaux, Villenave Dornon, France ; INRA Villenave Dornon, France .
    Sköllermo, Anna
    KTH, School of Biotechnology (BIO), Proteomics. KTH, School of Biotechnology (BIO), Centres, Albanova VinnExcellence Center for Protein Technology, ProNova.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics. KTH, School of Biotechnology (BIO), Centres, Albanova VinnExcellence Center for Protein Technology, ProNova.
    Blanchard, Alain
    Univ Bordeaux, Villenave Dornon, France ; INRA Villenave Dornon, France .
    Sirand-Pugnet, Pascal
    Univ Bordeaux, Villenave Dornon, France ; INRA Villenave Dornon, France .
    Specific Evolution of F-1-Like ATPases in Mycoplasmas2012In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, no 6, p. e38793-Article in journal (Refereed)
    Abstract [en]

    F1F0 ATPases have been identified in most bacteria, including mycoplasmas which have very small genomes associated with a host-dependent lifestyle. In addition to the typical operon of eight genes encoding genuine F1F0 ATPase (Type 1), we identified related clusters of seven genes in many mycoplasma species. Four of the encoded proteins have predicted structures similar to the alpha, beta, gamma and e subunits of F-1 ATPases and could form an F-1-like ATPase. The other three proteins display no similarity to any other known proteins. Two of these proteins are probably located in the membrane, as they have three and twelve predicted transmembrane helices. Phylogenomic studies identified two types of F-1-like ATPase clusters, Type 2 and Type 3, characterized by a rapid evolution of sequences with the conservation of structural features. Clusters encoding Type 2 and Type 3 ATPases were assumed to originate from the Hominis group of mycoplasmas. We suggest that Type 3 ATPase clusters may spread to other phylogenetic groups by horizontal gene transfer between mycoplasmas in the same host, based on phylogeny and genomic context. Functional analyses in the ruminant pathogen Mycoplasma mycoides subsp. mycoides showed that the Type 3 cluster genes were organized into an operon. Proteomic analyses demonstrated that the seven encoded proteins were produced during growth in axenic media. Mutagenesis and complementation studies demonstrated an association of the Type 3 cluster with a major ATPase activity of membrane fractions. Thus, despite their tendency toward genome reduction, mycoplasmas have evolved and exchanged specific F-1-like ATPases with no known equivalent in other bacteria. We propose a model, in which the F-1-like structure is associated with a hypothetical X-0 sector located in the membrane of mycoplasma cells.

  • 6.
    Gantelius, Jesper
    et al.
    KTH, School of Biotechnology (BIO), Nano Biotechnology.
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics.
    Hamsten, Carl
    KTH, School of Biotechnology (BIO).
    Neiman, Maja
    KTH, School of Biotechnology (BIO), Proteomics.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Andersson-Svahn, Helene
    KTH, School of Biotechnology (BIO).
    A lateral flow protein microarray for rapid determination of contagious bovine pleuropneumonia status in bovine serum2010In: Journal of Microbiological Methods, ISSN 0167-7012, E-ISSN 1872-8359, Vol. 82, no 1, p. 11-18Article in journal (Refereed)
    Abstract [en]

    Novel analytical methods for a next generation of diagnostic devices combine attributes from sensitive, accurate, fast, simple and multiplexed analysis methods. Here, we describe a possible contribution to these by the application of a lateral flow microarray where a panel of recombinant protein antigens was used to differentiate bovine serum samples in the context of the lung disease contagious bovine pleuropneumonia (CBPP). Lateral flow arrays were produced by attaching nitrocellulose onto microscopic slides and spotting of the recombinant proteins onto the membranes. The developed assay included evaluations of substrate matrix and detection reagents to allow for short assay times and convenient read-out options, and to yield a total assay time from sample application to data acquisition of less than ten minutes. It was found that healthy and disease-affected animals could be discriminated (AUC = 97%), and we suggest that the use of an antigen panel in combination with the lateral flow device offers an emerging analytical tool towards a simplified but accurate on-site diagnosis.

  • 7. Gaurivaud, P.
    et al.
    Persson, Anja
    KTH, Superseded Departments, Biotechnology.
    Le Grand, D.
    Westberg, Joakim
    KTH, Superseded Departments, Biotechnology.
    Solsona, M.
    Johansson, K. E.
    Poumarat, F.
    Variability of a glucose phosphotransferase system permease in Mycoplasma mycoides subsp mycoides Small Colony2004In: Microbiology, ISSN 1350-0872, E-ISSN 1465-2080, Vol. 150, p. 4009-4022Article in journal (Refereed)
    Abstract [en]

    Intraclonal antigenic variation in pathogenic mycoplasma species is considered an important feature of host-pathogen interaction. Such intraclonal protein variation was observed for the interaction of Mycoplasma mycoides subsp. mycoides Small Colony, the agent of contagious bovine pleuropneumonia, with mAb 3F3. Colony immunostaining allows the definition of 3F3 ON- and 3F3 OFF-type variants, which revert at low frequency. Targets of mAb 3F3 were shown to be surface located, and resided on multiple polypeptides in the 58-68 kDa size range. Phage display and a genomic database were combined to determine the gene encoding the proteins recognized by mAb 3F3. A gene encoding the putative permease of the glucose phosphotransferase system was identified. Genome sequence analysis of strain PG1 revealed two highly similar copies of this gene, resulting from duplication of the chromosomal region carrying the gene. Southern blot analysis demonstrated the presence of this duplication in almost every African strain tested, but not in European strains. DNA analysis revealed that ON/OFF switching is governed by a base substitution occurring upstream of the coding region for the 3F3 epitope. This event generates a stop codon that results in the premature termination of the PtsG protein.

  • 8.
    Hamsten, Carl
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Neiman, Maja
    KTH, School of Biotechnology (BIO), Proteomics.
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics.
    Hamsten, Marica
    KTH, School of Biotechnology (BIO), Proteomics.
    March, John B.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Recombinant surface proteomics as a tool to analyze humoral immune responses in bovines infected by Mycoplasma mycoides subsp. mycoides SC2009In: Molecular & Cellular Proteomics, ISSN 1535-9476, E-ISSN 1535-9484, Vol. 8, no 11, p. 2544-2554Article in journal (Refereed)
    Abstract [en]

    A systematic approach to characterize the surface proteome of Mycoplasma mycoides subspecies mycoides small colony type (M. mycoides SC), the causing agent of contagious bovine pleuropneumonia (CBPP) in cattle, is presented. Humoral immune responses in 242 CBPP affected cattle and controls were monitored against one third of the surface proteins of M. mycoides SC in a high-throughput magnetic bead based assay. First, 64 surface proteins were selected from the genome sequence of M. mycoides SC and expressed as recombinant proteins in E. coli. Binding of antibodies to each individual protein could then be analyzed simultaneously in minute sample volumes with the Luminex suspension array technology. The assay was optimized on Namibian CBPP positive sera and Swedish negative controls to allow detection and 20-fold mean signal separation between CBPP positive and negative sera. Signals were proven to be protein-specific by inhibition experiments and results agreed with western blot experiments. The assay's potential to monitor IgG, IgM and IgA responses over time was shown in a proof-of-concept study with 116 sera from 8 animals in a CBPP vaccine study. In conclusion, a toolbox with recombinant proteins and a flexible suspension array assay that allows multiplex analysis of humoral immune responses to M mycoides SC, has been created.

  • 9.
    Hamsten, Carl
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Tjipura-Zaire, Georgina
    McAuliffe, Laura
    Hübschle, Otto
    Scacchia, Massimo
    Ayling, Roger D.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Protein-Specific Analysis of Humoral Immune Responses in a Clinical Trial for Vaccines against Contagious Bovine Pleuropneumonia2010In: Clinical and Vaccine Immunology, ISSN 1556-6811, E-ISSN 1556-679X, Vol. 17, no 5, p. 853-861Article in journal (Refereed)
    Abstract [en]

    Specific humoral immune responses in a clinical trial on cattle for vaccines against contagious bovine pleuropneumonia (CBPP) were investigated. The trial included a subunit vaccine consisting of five recombinant putative variable surface proteins of the infectious agent Mycoplasma mycoides subspecies mycoides small colony type (M. mycoides SC) compared to the currently approved attenuated vaccine strain T1/44 and untreated controls. Humoral immune responses to 65 individual recombinant surface proteins of M. mycoides SC were monitored by a recently developed bead based array assay. Responses to the subunit vaccine components were found to be weak. Animals vaccinated with this vaccine were not protected and had CBPP lesions similar to the untreated controls. In correlating protein-specific humoral responses to T1/44 induced immunity, five proteins associated with a protective immune response were identified,namely LppQ and those of ORFs MSC_0271, MSC_0136, MSC_0079 and MSC_0431. The five proteins may be important candidates in the development of a novel subunit vaccine against CBPP.

  • 10.
    Hamsten, Carl
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Westberg, Joakim
    Bölske, Göran
    Ayling, Roger
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Expression and immunogenicity of six putative variable surface proteins in Mycoplasma mycoides subsp. mycoides SC.2008In: Microbiology, ISSN 1350-0872, E-ISSN 1465-2080, Vol. 154, p. 539-549Article in journal (Refereed)
    Abstract [en]

    Variable surface protein Vmm and five Vmm-type proteins from Mycoplasma mycoides subsp. mycoides SC were analysed to determine whether these proteins are expressed in vivo in animals affected by contagious bovine pleuropneumonia (CBPP) and in vitro. Recombinant versions of these proteins were constructed and expressed in Escherichia coli after mutation of the TGA Trp codons to TGG. These proteins were then analysed by dot and Western blotting with sera from CBPP-affected cattle. Furthermore, affinity-purified polyclonal antibodies to the recombinant proteins were used in Western and colony blotting to look for expression of the putative Vmm-type proteins in cultured M. mycoides SC. This study demonstrates that immunoglobulins in CBPP sera recognize all putative Vmm-type proteins tested, indicating that these proteins or their homologues are expressed by mycoplasmas during natural infections. Vmm and one of the putative Vmm-type proteins showed variable expression in vitro.

  • 11.
    Lindskog, Mats
    et al.
    KTH, School of Biotechnology (BIO).
    Berglund, L.
    Persson, Anja
    KTH, School of Biotechnology (BIO).
    Sivertsson, Åsa
    KTH, School of Biotechnology (BIO).
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO).
    Al-Khalili Szigyarto, Cristina
    KTH, School of Biotechnology (BIO).
    Design of protein epitope signature tags for antibody-based proteomicsManuscript (Other academic)
  • 12. Miles, K.
    et al.
    McAuliffe, L.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Ayling, R. D.
    Nicholas, R. A. J.
    Insertion sequence profiling of UK Mycoplasma bovis field isolates2005In: Veterinary Microbiology, ISSN 0378-1135, E-ISSN 1873-2542, Vol. 107, no 04-mar, p. 301-306Article in journal (Refereed)
    Abstract [en]

    The presence and distribution of insertion sequences ISMbov2 and ISMbov3 within Mycoplasma bovis were investigated. Analysis was carried out by Southern blotting using specific probes of 221 bp and 185 bp, to detect ISMbov2 and ISMbov3, respectively, amplified from the homologous sequences ISMmyl and ISI634 within Mycoplasma mycoides subspecies mycoides small colony type. We present data obtained from 49 field isolates of M. bovis, originating from pneumonic lungs, collected within the United Kingdom between 1996 and 2002. Hybridisation profiles show considerable variation between strains. ISMbov2 sequences are present between 2 and 17 copies while there are between 3 and 14 copies of the ISI634 homologue ISMbov3. These data also provide support for previous analysis by random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP). Crown

  • 13.
    Neiman, Maja
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Hamsten, Carl
    KTH, School of Biotechnology (BIO), Proteomics.
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics.
    Bölske, Göran
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Multiplex Screening of Surface Proteins from Mycoplasma mycoides subsp mycoides Small Colony for an Antigen Cocktail Enzyme-Linked Immunosorbent Assay2009In: Clinical and Vaccine Immunology, ISSN 1556-6811, E-ISSN 1556-679X, Vol. 16, no 11, p. 1665-1674Article in journal (Refereed)
    Abstract [en]

    A recombinant antigen cocktail ELISA for diagnosis of contagious bovine pleuropneumonia (CBPP) was developed after careful selection of antigens among one third of the surface proteome of the infectious agent Mycoplasma mycoides subsp. mycoides SC (M. mycoides SC). First, a miniaturized and parallelized assay system employing antigen suspension bead array technology was used to screen 97 bovine sera for their humoral immune response towards 61 recombinant surface proteins from M. mycoides SC. Statistical analysis of the data resulted in selection of eight proteins that showed strong serologic responses in CBPP-affected sera and minimal reactivity in negative control sera, with p-values less than 10-6. Only minor cross reactivity to hyperimmune sera against other mycoplasmas was observed. When applied in an ELISA, the cocktail of eight recombinant antigens allowed a five fold signal separation between 24 CBPP-affected and 23 CBPP-free sera from different geographical origin. No false positives and only two false negatives were obtained. In conclusion, the selected recombinant mycoplasma antigens qualified as highly specific markers for CBPP and could be employed in both a suspension bead array platform and a cocktail ELISA setting. This set of proteins and technologies therefore offer a powerful combination to drive and further improve serological assays towards reliable, simple and cost-effective diagnosis of CBPP.

  • 14.
    Nilsson, Peter
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Larsson, Karin
    KTH, School of Biotechnology (BIO).
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    Wernérus, Henrik
    KTH, School of Biotechnology (BIO).
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Proteomics.
    Björling, Erik
    KTH, School of Biotechnology (BIO).
    Ottoson, Jenny
    KTH, School of Biotechnology (BIO), Proteomics.
    Ödling, Jenny
    KTH, School of Biotechnology (BIO).
    Sundberg, Mårten
    KTH, School of Biotechnology (BIO), Proteomics.
    Al-Khalili Szigyarto, Cristina
    KTH, School of Biotechnology (BIO), Proteomics.
    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 profiling2005In: Proteomics, ISSN 1615-9853, E-ISSN 1615-9861, Vol. 5, p. 4327-4337Article in journal (Refereed)
    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.

  • 15.
    Persson, Anja
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Proteomics.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    A human protein atlas based on antibody proteomics2006In: Current opinion in molecular therapeutics (Print), ISSN 1464-8431, E-ISSN 2040-3445, Vol. 8, no 3, p. 185-190Article in journal (Refereed)
    Abstract [en]

    The Human Protein Atlas is a comprehensive database that provides the protein expression profiles for a large number of human proteins, presented as immunohistological images from most human tissues. This review provides an overview of the contents of the atlas, discusses the project strategy and highlights the importance of open access for data validation and quality. Essential procedures that are implemented during antibody production and image generation, such as the use of protein epitope signature tags ( PrEST) antigens, monospecific antibodies, tissue microarrays and thorough quality validation, are also discussed. The Human Protein Atlas is related to four other expression atlas initiatives, including, in particular, an upcoming protein atlas developed by the Sanger Institute.

  • 16. Ponten, Fredrik
    et al.
    Gry, Marcus
    KTH, School of Biotechnology (BIO), Proteomics.
    Fagerberg, Linn
    KTH, School of Biotechnology (BIO), Proteomics.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics.
    Asplund, Anna
    Berglund, Lisa
    KTH, School of Biotechnology (BIO), Proteomics.
    Oksvold, Per
    KTH, School of Biotechnology (BIO), Proteomics.
    Björling, Erik
    KTH, School of Biotechnology (BIO), Proteomics.
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Proteomics.
    Kampf, Caroline
    Navani, Sanjay
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics.
    Ottosson, Jenny
    KTH, School of Biotechnology (BIO), Proteomics.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Wernérus, Henrik
    KTH, School of Biotechnology (BIO), Proteomics.
    Wester, Kenneth
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    A global view of protein expression in human cells, tissues, and organs2009In: Molecular Systems Biology, ISSN 1744-4292, E-ISSN 1744-4292, Vol. 5Article in journal (Refereed)
    Abstract [en]

    Defining the protein profiles of tissues and organs is critical to understanding the unique characteristics of the various cell types in the human body. In this study, we report on an anatomically comprehensive analysis of 4842 protein profiles in 48 human tissues and 45 human cell lines. A detailed analysis of over 2 million manually annotated, high-resolution, immunohistochemistry- based images showed a high fraction (>65%) of expressed proteins in most cells and tissues, with very few proteins (<2%) detected in any single cell type. Similarly, confocal microscopy in three human cell lines detected expression of more than 70% of the analyzed proteins. Despite this ubiquitous expression, hierarchical clustering analysis, based on global protein expression patterns, shows that the analyzed cells can be still subdivided into groups according to the current concepts of histology and cellular differentiation. This study suggests that tissue specificity is achieved by precise regulation of protein levels in space and time, and that different tissues in the body acquire their unique characteristics by controlling not which proteins are expressed but how much of each is produced. Molecular Systems Biology 5: 337; published online 22 December 2009; doi:10.1038/msb.2009.93

  • 17.
    Pontén, Fredrik
    et al.
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University.
    Gry, Marcus
    KTH, School of Biotechnology (BIO), Proteomics.
    Björling, Erik
    KTH, School of Biotechnology (BIO), Proteomics.
    Berglund, Lisa
    KTH, School of Biotechnology (BIO), Proteomics.
    Al-Khalili Szigyarto, Cristina
    KTH, School of Biotechnology (BIO), Proteomics.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics.
    Andersson-Svahn, Helene
    KTH, School of Biotechnology (BIO), Proteomics.
    Asplund, Anna
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University.
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Proteomics.
    Kampf, Caroline
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University.
    Nilsson, Kenneth
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics.
    Ottosson, Jenny
    KTH, School of Biotechnology (BIO), Proteomics.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Wernérus, Henrik
    KTH, School of Biotechnology (BIO), Proteomics.
    Wester, Kenneth
    Department of Genetics and Pathology, The Rudbeck Laboratory, Uppsala University.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    Ubiquitous protein expression in human cells, tissues and organsManuscript (Other academic)
  • 18. Schieck, Elise
    et al.
    Liljander, Anne
    Hamsten, Carl
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Gicheru, Nimmo
    Scacchia, Massimo
    Sacchini, Flavio
    Heller, Martin
    Schnee, Christiane
    Sterner-Kock, Anja
    Hlinak, Andreas
    Naessens, Jan
    Poole, Jane
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Jores, Joerg
    High antibody titres against predicted Mycoplasma surface proteins do not prevent sequestration in infected lung tissue in the course of experimental contagious bovine pleuropneumonia2014In: Veterinary Microbiology, ISSN 0378-1135, E-ISSN 1873-2542, Vol. 172, no 1-2, p. 285-293Article in journal (Refereed)
    Abstract [en]

    Contagious bovine pleuropneumonia (CBPP), a severe respiratory disease of cattle caused by Mycoplasma mycoides subsp. mycoides (Mmm) is endemic in many African countries due to fragmented veterinary services and the lack of an efficient vaccine and sensitive diagnostics. More efficient tools to control the disease are needed, but to develop the tools, a better understanding of host-pathogen interactions is necessary. The aim of this study was to characterize the kinetics of the humoral immune response against 65 Mmm surface antigens for an extended period in cattle that survived a primary infection with Mmm. We describe clinical and haematological outcomes, and dissect the humoral immune response over time, to specific antigens and compared the antibody responses between different pathomorphological outcomes. No antigen-specific antibodies correlating with protection were identified. Interestingly we found that animals that developed MycopIasma-containing sequestra had significantly higher antibody levels against proteins comprising the surface proteome than the animals that cleared Mycoplasma from their lungs. Based on these data we suggest that high antibody titres might play a role in the establishment of pathomorphological changes, such as vasculitis, which should be investigated in future studies. Beneficial antibody specificities and cellular immune responses need to be identified in order to foster the development of an improved vaccine in the future.

  • 19.
    Strömberg, Sara
    et al.
    Department of Genetics and Pathology, Rudbeck Laboratory, Uppsala University.
    Gry Björklund, Marcus
    KTH, School of Biotechnology (BIO), Proteomics.
    Asplund, Caroline
    KTH, School of Biotechnology (BIO), Proteomics.
    Sköllermo, Anna
    KTH, School of Biotechnology (BIO), Proteomics.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Wester, Kenneth
    Department of Genetics and Pathology, Rudbeck Laboratory, Uppsala University.
    Kampf, Caroline
    Department of Genetics and Pathology, Rudbeck Laboratory, Uppsala University.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics.
    Andersson, Ann-Catrin
    Department of Genetics and Pathology, Rudbeck Laboratory, Uppsala University.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    Kononen, Juha
    Beecher Instruments, Sun Prairie, WI, United States.
    Pontén, Fredrik
    Department of Genetics and Pathology, Rudbeck Laboratory, Uppsala University.
    Asplund, Anna
    Department of Genetics and Pathology, Rudbeck Laboratory, Uppsala University.
    A high-throughput strategy for protein profiling in cell microarrays using automated image analysis2007In: Proteomics, ISSN 1615-9853, E-ISSN 1615-9861, Vol. 7, no 13, p. 2142-2150Article in journal (Refereed)
    Abstract [en]

    Advances in antibody production render a growing supply of affinity reagents for immunohistochemistry (IHC), and tissue microarray (TMA) technologies facilitate simultaneous analysis of protein expression in a multitude of tissues. However, collecting validated IHC data remains a bottleneck problem, as the standard method is manual microscopical analysis. Here we present a high-throughput strategy combining IHC on a recently developed cell microarray with a novel, automated image-analysis application (TMAx). The software was evaluated on 200 digital images of IHC-stained cell spots, by comparing TMAx annotation with manual annotation performed by seven human experts. A high concordance between automated and manual annotation of staining intensity and fraction of IHC-positive cells was found. in a limited study, we also investigated the possibility to assess the correlation between mRNA and protein levels, by using TMAx output results for relative protein quantification and quantitative real-time PCR for the quantification of corresponding transcript levels. In conclusion, automated analysis of immunohistochemically stained in vitro-cultured cells in a microarray format can be used for high-throughput protein profiling, and extraction of RNA from the same cell lines provides a basis for comparing transcription and protein expression on a global scale.

  • 20.
    Tegel, Hanna
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Tourle, Samuel
    KTH, School of Biotechnology (BIO), Proteomics.
    Ottosson, Jenny
    KTH, School of Biotechnology (BIO), Proteomics.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Increased levels of recombinant human proteins with the Escherichia coli strain Rosetta(DE3)2010In: Protein Expression and Purification, ISSN 1046-5928, E-ISSN 1096-0279, Vol. 69, no 2, p. 159-167Article in journal (Refereed)
    Abstract [en]

    The effect of two Escherichia coli expression strains on the production of recombinant human protein fragments was evaluated. High-throughput protein production projects, such as the Swedish Human Protein Atlas project, are dependent on high protein yield and purity. By changing strain from E. coli BL21(DE3) to E. coli Rosetta(DE3) the overall success rate of the protein production has increased dramatically. The Rosetta(DE3) strain compensates for a number of rare codons. Here, we describe how the protein expression of human gene fragments in E. coli strains BL21(DE3) and Rosetta(DE3) was evaluated in two stages. Initially a test set of 68 recombinant proteins that previously had been expressed in BL21(DE3) was retransformed and expressed in Rosetta(DE3). The test set generated very positive results with an improved expression yield and a significantly better purity of the protein product which prompted us to implement the Rosetta(DE3) strain in the high-throughput protein production. Except for analysis of protein yield and purity the sequences were also analyzed regarding number of rare codons and rare codon clusters. The content of rare codons showed to have a significant effect on the protein purity. Based on the results of this study the atlas project permanently changed expression strain to Rosetta(DE3).

  • 21.
    Uhlén, Mathias
    et al.
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Björling, Erik
    KTH, School of Biotechnology (BIO).
    Agaton, Charlotta
    KTH, School of Biotechnology (BIO).
    Al-Khalili Szigyarto, Cristina
    KTH, School of Biotechnology (BIO).
    Amini, Bahram
    KTH, School of Biotechnology (BIO).
    Andersen, Elisabet
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Andersson, Ann-Catrin
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Angelidou, Pia
    KTH, School of Biotechnology (BIO).
    Asplund, Anna
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Asplund, Caroline
    KTH, School of Biotechnology (BIO).
    Berglund, Lisa
    KTH, School of Biotechnology (BIO).
    Bergström, Kristina
    KTH, School of Biotechnology (BIO).
    Brumer, Harry
    KTH, School of Biotechnology (BIO).
    Cerjan, Dijana
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Ekström, Marica
    KTH, School of Biotechnology (BIO).
    Elobeid, Adila
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Eriksson, Cecilia
    KTH, School of Biotechnology (BIO).
    Fagerberg, Linn
    KTH, School of Biotechnology (BIO).
    Falk, Ronny
    KTH, School of Biotechnology (BIO).
    Fall, Jenny
    KTH, School of Biotechnology (BIO).
    Forsberg, Mattias
    KTH, School of Biotechnology (BIO).
    Gry Björklund, Marcus
    KTH, School of Biotechnology (BIO).
    Gumbel, Kristoffer
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Halimi, Asif
    KTH, School of Biotechnology (BIO).
    Hallin, Inga
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Hamsten, Carl
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Hansson, Marianne
    KTH, School of Biotechnology (BIO).
    Hedhammar, My
    KTH, School of Biotechnology (BIO).
    Hercules, Görel
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Kampf, Caroline
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Larsson, Karin
    KTH, School of Biotechnology (BIO).
    Lindskog, Mats
    KTH, School of Biotechnology (BIO).
    Lodewyckx, Wald
    KTH, School of Biotechnology (BIO).
    Lund, Jan
    KTH, School of Biotechnology (BIO).
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO).
    Magnusson, Kristina
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Malm, Erik
    KTH, School of Biotechnology (BIO).
    Nilsson, Peter
    KTH, School of Biotechnology (BIO).
    Ödling, Jenny
    KTH, School of Biotechnology (BIO).
    Oksvold, Per
    KTH, School of Biotechnology (BIO).
    Olsson, Ingmarie
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Öster, Emma
    KTH, School of Biotechnology (BIO).
    Ottosson, Jenny
    KTH, School of Biotechnology (BIO).
    Paavilainen, Linda
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Rimini, Rebecca
    KTH, School of Biotechnology (BIO).
    Rockberg, Johan
    KTH, School of Biotechnology (BIO).
    Runeson, Marcus
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Sivertsson, Åsa
    KTH, School of Biotechnology (BIO).
    Sköllermo, Anna
    KTH, School of Biotechnology (BIO).
    Steen, Johanna
    KTH, School of Biotechnology (BIO).
    Stenvall, Maria
    KTH, School of Biotechnology (BIO).
    Sterky, Fredrik
    KTH, School of Biotechnology (BIO).
    Strömberg, Sara
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Sundberg, Mårten
    KTH, School of Biotechnology (BIO).
    Tegel, Hanna
    KTH, School of Biotechnology (BIO).
    Tourle, Samuel
    KTH, School of Biotechnology (BIO).
    Wahlund, Eva
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Waldén, Annelie
    KTH, School of Biotechnology (BIO).
    Wan, Jinghong
    KTH, School of Biotechnology (BIO), Molecular Biotechnology (closed 20130101).
    Wernérus, Henrik
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Westberg, Joakim
    KTH, School of Biotechnology (BIO).
    Wester, Kenneth
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    Wrethagen, Ulla
    KTH, School of Biotechnology (BIO).
    Xu, Lan Lan
    KTH, School of Biotechnology (BIO).
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Pontén, Fredrik
    Uppsala Univ, Rudbeck Lab, Dept Genet & Pathol.
    A human protein atlas for normal and cancer tissues based on antibody proteomics2005In: Molecular & Cellular Proteomics, ISSN 1535-9476, E-ISSN 1535-9484, Vol. 4, no 12, p. 1920-1932Article in journal (Refereed)
    Abstract [en]

    Antibody-based proteomics provides a powerful approach for the functional study of the human proteome involving the systematic generation of protein-specific affinity reagents. We used this strategy to construct a comprehensive, antibody-based protein atlas for expression and localization profiles in 48 normal human tissues and 20 different cancers. Here we report a new publicly available database containing, in the first version, similar to 400,000 high resolution images corresponding to more than 700 antibodies toward human proteins. Each image has been annotated by a certified pathologist to provide a knowledge base for functional studies and to allow queries about protein profiles in normal and disease tissues. Our results suggest it should be possible to extend this analysis to the majority of all human proteins thus providing a valuable tool for medical and biological research.

  • 22. Vashisht, Ajay A.
    et al.
    Zumbrennen, Kimberly B.
    Huang, Xinhua
    Powers, David N.
    Durazo, Armando
    Sun, Dahui
    Bhaskaran, Nimesh
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    Sangfelt, Olle
    Spruck, Charles
    Leibold, Elizabeth A.
    Wohlschlegel, James A.
    Control of Iron Homeostasis by an Iron-Regulated Ubiquitin Ligase2009In: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 326, no 5953, p. 718-721Article in journal (Refereed)
    Abstract [en]

    Eukaryotic cells require iron for survival and have developed regulatory mechanisms for maintaining appropriate intracellular iron concentrations. The degradation of iron regulatory protein 2 (IRP2) in iron-replete cells is a key event in this pathway, but the E3 ubiquitin ligase responsible for its proteolysis has remained elusive. We found that a SKP1-CUL1-FBXL5 ubiquitin ligase protein complex associates with and promotes the iron-dependent ubiquitination and degradation of IRP2. The F-box substrate adaptor protein FBXL5 was degraded upon iron and oxygen depletion in a process that required an iron-binding hemerythrin-like domain in its N terminus. Thus, iron homeostasis is regulated by a proteolytic pathway that couples IRP2 degradation to intracellular iron levels through the stability and activity of FBXL5.

  • 23. Westberg, J.
    et al.
    Persson, Anja
    Pettersson, B.
    Uhlén, Mathias
    KTH, Superseded Departments, Biotechnology.
    Johansson, K. E.
    ISMmy1, a novel insertion sequence of Mycoplasma mycoides subsp mycoides small colony type2002In: FEMS Microbiology Letters, ISSN 0378-1097, E-ISSN 1574-6968, Vol. 208, no 2, p. 207-213Article in journal (Refereed)
    Abstract [en]

    A new insertion sequence, ISMmy1, has been identified in the bovine pathogen Mycoplasma mycoides subsp. mycoides biotype small colony MmymySC. The occurrence of ISMmy1 in 15 MmymySC strains and 12 other mycoplasmas was examined by Southern blotting. All MmymySC strains showed identical hybridisation patterns except for the type strain PG11, the vaccine strain T1Sr49, and the strain Afade, which all had unique patterns. ISMmy1-like sequences were also found in the bovine pathogen Mycoplasma bovis strain Donetta(T) while mycoplasmas that are phylogenetically closer to MmymySC lack ISMmy1. This observation suggests horizontal transfer between MmymySC and M. bovis.

  • 24.
    Westberg, Joakim
    et al.
    KTH, Superseded Departments, Biotechnology.
    Persson, Anja
    KTH, Superseded Departments, Biotechnology.
    Holmberg, Anders H.
    KTH, Superseded Departments, Biotechnology.
    Goesmann, A.
    Lundeberg, Joakim
    KTH, Superseded Departments, Biotechnology.
    Johansson, K. E.
    Pettersson, Bertil
    KTH, Superseded Departments, Biotechnology.
    Uhlén, Mathias
    KTH, Superseded Departments, Biotechnology.
    The genome sequence of Mycoplasma mycoides subsp mycoides SC type strain PG1(T), the causative agent of contagious bovine pleuropneumonia (CBPP)2004In: Genome Research, ISSN 1088-9051, E-ISSN 1549-5469, Vol. 14, no 2, p. 221-227Article in journal (Refereed)
    Abstract [en]

    Mycoplasma mycoides subsp. mycoidesSC (MmymySC) is the etiological agent of contagious bovine pleuropneumonia (CBPP), a highly contagious respiratory disease in cattle. The genome of Mmymy SC type strain PUT has been sequenced to map all the genes and to facilitate further studies regarding the cell function of the organism and CBPP. The genome is characterized by a single circular chromosome of 1,211,703 bp with the lowest G+C content (24 mole%) and the highest density of insertion sequences (13% of the genome size) of all sequenced bacterial genomes. The genome contains 985 putative genes, of which 72 are part of insertion sequences and encode transposases. Anomalies in the GC-skew pattern and the presence of large repetitive sequences indicate a high genomic plasticity. A variety of potential virulence factors was identified, including genes encoding putative variable surface proteins and enzymes and transport proteins responsible for the production of hydrogen peroxide and the capsule, which is believed to have toxic effects on the animal.

  • 25.
    Älgenäs, Cajsa
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Agaton, Charlotta
    Fagerberg, Linn
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Asplund, Anna
    Björling, Lisa
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Björling, Erik
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Kampf, Caroline
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Persson, Anja
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Wester, Kenneth
    Pontén, Fredrik
    Wernerus, Henrik
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Ottosson Takanen, Jenny
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Protein Technology.
    Antibody performance in western blot applications is context- dependent2014In: Biotechnology Journal, ISSN 1860-6768, E-ISSN 1860-7314, Vol. 9, no 3, p. 435-445Article in journal (Refereed)
    Abstract [en]

    An important concern for the use of antibodies in various applications, such as western blot (WB) or immunohistochemistry (IHC), is specificity. This calls for systematic validations using well-designed conditions. Here, we have analyzed 13000 antibodies using western blot with lysates from human cell lines, tissues, and plasma. Standardized stratification showed that 45% of the antibodies yielded supportive staining, and the rest either no staining (12%) or protein bands of wrong size (43%). A comparative study of WB and IHC showed that the performance of antibodies is application-specific, although a correlation between no WB staining and weak IHC staining could be seen. To investigate the influence of protein abundance on the apparent specificity of the antibody, new WB analyses were performed for 1369 genes that gave unsupportive WBs in the initial screening using cell lysates with overexpressed full-length proteins. Then, more than 82% of the antibodies yielded a specific band corresponding to the full-length protein. Hence, the vast majority of the antibodies (90%) used in this study specifically recognize the target protein when present at sufficiently high levels. This demonstrates the context- and application-dependence of antibody validation and emphasizes that caution is needed when annotating binding reagents as specific or cross-reactive. WB is one of the most commonly used methods for validation of antibodies. Our data implicate that solely using one platform for antibody validation might give misleading information and therefore at least one additional method should be used to verify the achieved data.

1 - 25 of 25
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