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  • 1.
    Ayoglu, Burcu
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Mitsios, Nicholas
    Kockum, Ingrid
    Khademi, Mohsen
    Zandian, Arash
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sjoberg, Ronald
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Forsstrom, Bjorn
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Bredenberg, Johan
    Bomfim, Izaura Lima
    Holmgren, Erik
    Gronlund, Hans
    Guerreiro-Cacais, Andre Ortlieb
    Abdelmagid, Nada
    Uhlen, Mathias
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Waterboer, Tim
    Alfredsson, Lars
    Mulder, Jan
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Olsson, Tomas
    Nilsson, Peter
    Anoctamin 2 identified as an autoimmune target in multiple sclerosis2016In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 113, no 8, p. 2188-2193Article in journal (Refereed)
    Abstract [en]

    Multiple sclerosis (MS) is the most common chronic inflammatory disease of the central nervous system and also is regarded as an autoimmune condition. However, the antigenic targets of the autoimmune response in MS have not yet been deciphered. In an effort to mine the autoantibody repertoire within MS, we profiled 2,169 plasma samples from MS cases and population-based controls using bead arrays built with 384 human protein fragments selected from an initial screening with 11,520 antigens. Our data revealed prominently increased autoantibody reactivity against the chloride-channel protein anoctamin 2 (ANO2) in MS cases compared with controls. This finding was corroborated in independent assays with alternative protein constructs and by epitope mapping with peptides covering the identified region of ANO2. Additionally, we found a strong interaction between the presence of ANO2 autoantibodies and the HLA complex MS-associated DRB1*15 allele, reinforcing a potential role for ANO2 autoreactivity in MS etiopathogenesis. Furthermore, immunofluorescence analysis in human MS brain tissue showed ANO2 expression as small cellular aggregates near and inside MS lesions. Thus this study represents one of the largest efforts to characterize the autoantibody repertoire within MS. The findings presented here demonstrate that an ANO2 autoimmune subphenotype may exist in MS and lay the groundwork for further studies focusing on the pathogenic role of ANO2 autoantibodies in MS.

  • 2.
    Ayoglu, Burcu
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sjöberg, Ronald
    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.
    Schwenk, Jochen M.
    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.
    et al.,
    The calcium-activated chloride channel anoctamine 2 as an autoimmune component of multiple sclerosisManuscript (preprint) (Other academic)
  • 3.
    Bremer, Hanna D.
    et al.
    Swedish Univ Agr Sci, Dept Clin Sci, SE-75007 Uppsala, Sweden..
    Landegren, Nils
    Karolinska Inst, Karolinska Univ Hosp, Dept Med Solna, CMM, L8 01, SE-17176 Stockholm, Sweden.;Uppsala Univ, Dept Med Sci, Sci Life Lab, Uppsala, Sweden..
    Sjöberg, Ronald
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hallgren, Asa
    Karolinska Inst, Karolinska Univ Hosp, Dept Med Solna, CMM, L8 01, SE-17176 Stockholm, Sweden..
    Renneker, Stefanie
    Euroimmun AG, D-23560 Lubeck, Germany..
    Lattwein, Erik
    Euroimmun AG, D-23560 Lubeck, Germany..
    Leonard, Dag
    Uppsala Univ, Rheumatol & Sci Life Lab, Dept Med Sci, SE-75185 Uppsala, Sweden..
    Eloranta, Maija-Leena
    Uppsala Univ, Rheumatol & Sci Life Lab, Dept Med Sci, SE-75185 Uppsala, Sweden..
    Ronnblom, Lars
    Uppsala Univ, Rheumatol & Sci Life Lab, Dept Med Sci, SE-75185 Uppsala, Sweden..
    Nordmark, Gunnel
    Uppsala Univ, Rheumatol & Sci Life Lab, Dept Med Sci, SE-75185 Uppsala, Sweden..
    Nilsson, Peter
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics.
    Andersson, Goran
    Swedish Univ Agr Sci, Dept Anim Breeding & Genet, SE-75007 Uppsala, Sweden..
    Lilliehook, Inger
    Swedish Univ Agr Sci, Dept Clin Sci, SE-75007 Uppsala, Sweden..
    Lindblad-Toh, Kerstin
    Broad Inst Harvard & MIT, Cambridge, MA 02142 USA.;Uppsala Univ, Sci Life Lab, IMBIM, SE-75123 Uppsala, Sweden..
    Kampe, Olle
    Karolinska Inst, Karolinska Univ Hosp, Dept Med Solna, CMM, L8 01, SE-17176 Stockholm, Sweden.;Uppsala Univ, Dept Med Sci, Sci Life Lab, Uppsala, Sweden.;Univ Bergen, Dept Clin Sci, N-5021 Bergen, Norway.;Univ Bergen, KG Jebsen Ctr Autoimmune Disorders, N-5021 Bergen, Norway.;Haukeland Hosp, Dept Med, N-5021 Bergen, Norway..
    Hansson-Hamlin, Helene
    Swedish Univ Agr Sci, Dept Clin Sci, SE-75007 Uppsala, Sweden..
    ILF2 and ILF3 are autoantigens in canine systemic autoimmune disease2018In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 8, article id 4852Article in journal (Refereed)
    Abstract [en]

    Dogs can spontaneously develop complex systemic autoimmune disorders, with similarities to human autoimmune disease. Autoantibodies directed at self-antigens are a key feature of these autoimmune diseases. Here we report the identification of interleukin enhancer-binding factors 2 and 3 (ILF2 and ILF3) as autoantigens in canine immune-mediated rheumatic disease. The ILF2 autoantibodies were discovered in a small, selected canine cohort through the use of human protein arrays; a method not previously described in dogs. Subsequently, ILF3 autoantibodies were also identified in the same cohort. The results were validated with an independent method in a larger cohort of dogs. ILF2 and ILF3 autoantibodies were found exclusively, and at a high frequency, in dogs that showed a speckled pattern of antinuclear antibodies on immunofluorescence. ILF2 and ILF3 autoantibodies were also found at low frequency in human patients with SLE and Sjogren's syndrome. These autoantibodies have the potential to be used as diagnostic biomarkers for canine, and possibly also human, autoimmune disease.

  • 4. Colwill, Karen
    et al.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics.
    Sundberg, Mårten
    KTH, School of Biotechnology (BIO), Proteomics.
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics.
    Sivertsson, Åsa
    KTH, School of Biotechnology (BIO), Proteomics.
    Schwenk, Jochen M
    KTH, School of Biotechnology (BIO), Proteomics.
    Ottosson Takanen, Jenny
    KTH, School of Biotechnology (BIO), Proteomics.
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Proteomics.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    Gräslund, Susanne
    et, al.
    A roadmap to generate renewable protein binders to the human proteome2011In: Nature Methods, ISSN 1548-7091, E-ISSN 1548-7105, Vol. 8, no 7, p. 551-8Article in journal (Refereed)
    Abstract [en]

    Despite the wealth of commercially available antibodies to human proteins, research is often hindered by their inconsistent validation, their poor performance and the inadequate coverage of the proteome. These issues could be addressed by systematic, genome-wide efforts to generate and validate renewable protein binders. We report a multicenter study to assess the potential of hybridoma and phage-display technologies in a coordinated large-scale antibody generation and validation effort. We produced over 1,000 antibodies targeting 20 SH2 domain proteins and evaluated them for potency and specificity by enzyme-linked immunosorbent assay (ELISA), protein microarray and surface plasmon resonance (SPR). We also tested selected antibodies in immunoprecipitation, immunoblotting and immunofluorescence assays. Our results show that high-affinity, high-specificity renewable antibodies generated by different technologies can be produced quickly and efficiently. We believe that this work serves as a foundation and template for future larger-scale studies to create renewable protein binders.

  • 5.
    Gantelius, Jesper
    et al.
    KTH, School of Biotechnology (BIO), Nano Biotechnology (closed 20130101).
    Bass, Tarek
    KTH, School of Biotechnology (BIO), Molecular Biotechnology (closed 20130101).
    Gundberg, Anna
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Sundberg, Mårten
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Andersson-Svahn, Helene
    KTH, School of Biotechnology (BIO), Nano Biotechnology (closed 20130101).
    A ten-minute high density lateral flow protein microarray assay2011In: 15th International Conference on Miniaturized Systems for Chemistry and Life Sciences 2011: (MicroTAS 2011), 2011, p. 1176-1178Conference paper (Refereed)
    Abstract [en]

    Protein microarrays are useful tools for highly multiplexed determination of presence or levels of clinically relevant biomarkers in human tissues and biofluids. However, such tools have thus far been restricted to laboratory environments. Here, we present a novel 384-plexed easy to use lateral flow protein microarray device capable of sensitive (<50ng/ml) determination of antigen specific antibodies in less than ten minutes total assay time. Results were developed with gold nanobeads and could be recorded by a cell-phone camera or table top scanner. Excellent accuracy (AUC=99.4%) was achieved in comparison with an established glass microarray assay for 26 antigen-specific antibodies.

  • 6.
    Gantelius, Jesper
    et al.
    KTH, School of Biotechnology (BIO), Nano Biotechnology (closed 20130101).
    Bass, Tarek
    KTH, School of Biotechnology (BIO), Molecular Biotechnology (closed 20130101).
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Andersson-Svahn, Helene
    KTH, School of Biotechnology (BIO), Nano Biotechnology (closed 20130101).
    A Lateral Flow Protein Microarray for Rapid and Sensitive Antibody Assays2011In: International Journal of Molecular Sciences, ISSN 1661-6596, Vol. 12, no 11, p. 7748-7759Article in journal (Refereed)
    Abstract [en]

    Protein microarrays are useful tools for highly multiplexed determination of presence or levels of clinically relevant biomarkers in human tissues and biofluids. However, such tools have thus far been restricted to laboratory environments. Here, we present a novel 384-plexed easy to use lateral flow protein microarray device capable of sensitive (<30 ng/mL) determination of antigen-specific antibodies in ten minutes of total assay time. Results were developed with gold nanobeads and could be recorded by a cell-phone camera or table top scanner. Excellent accuracy with an area under curve (AUC of 98% was achieved in comparison with an established glass microarray assay for 26 antigen-specific antibodies. We propose that the presented framework could find use in convenient and cost-efficient quality control of antibody production, as well as in providing a platform for multiplexed affinity-based assays in low-resource or mobile settings.

  • 7.
    Hellström, Cecilia
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Dodig-Crnković, Tea
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hong, Mun-Gwan
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Schwenk, Jochen
    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.
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    High-density serum/plasma reverse phase protein arrays2017In: Serum/Plasma Proteomics, Humana Press, 2017, p. 229-238Chapter in book (Refereed)
    Abstract [en]

    In-depth exploration and characterization of human serum and plasma proteomes is an attractive strategy for the identification of potential prognostic or diagnostic biomarkers. The possibility of analyzing larger numbers of samples in a high-throughput fashion has markedly increased with affinity-based microarrays, thus providing higher statistical power to these biomarker studies. Here, we describe a protocol for high-density serum and plasma reverse phase protein arrays (RPPAs). We demonstrate how a biobank of 12,392 samples was immobilized and analyzed on a single microarray slide, allowing high-quality profiling of abundant target proteins across all samples in one assay.

  • 8. Janzi, Magdalena
    et al.
    Kull, Inger
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics.
    Wan, Jinghong
    KTH, School of Biotechnology (BIO), Proteomics.
    Melen, Erik
    Bayat, Narges
    Ostblom, Eva
    Pan-Hammarstrom, Qiang
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics.
    Hammarstrom, Lennart
    Selective IgA deficiency in early life: Association to infections and allergic diseases during childhood2009In: Clinical Immunology, ISSN 1521-6616, E-ISSN 1521-7035, Vol. 133, no 1, p. 78-85Article in journal (Refereed)
    Abstract [en]

    Selective IgA deficiency in early life is quite common in Caucasian populations, but it is unclear whether it increases the risk of infections and allergic diseases during childhood. Serum IgA levels were measured in 2423 children at 4 years of age in a Swedish population based birth cohort (BAMSE). Parental questionnaires were repeatedly sent out during the child's first 8 years of life, collecting information about infections and allergic diseases. 14 children (1:173) were found to be IgA deficient at 4 years of age. These children had an increased risk of pseudocroup at year 1 (p<0.01) and food hypersensitivity at year 4 (p<0.05) as compared to IgA sufficient children. No increased risk was observed in the partial IgA deficiency group. The findings suggest that selective IgA deficiency may increase the risk of parentally reported pseudocroup and food hypersensitivity during early childhood.

  • 9. Janzi, Magdalena
    et al.
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Wan, Jinghong
    Fischler, Björn
    von Döbeln, Ulrika
    Isaac, Lourdes
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hammarström, Lennart
    Screening for C3 deficiency in newborns using microarrays.2009In: PloS one, ISSN 1932-6203, Vol. 4, no 4, p. e5321-Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Dried blood spot samples (DBSS) from newborns are widely used in neonatal screening for selected metabolic diseases and diagnostic possibilities for additional disorders are continuously being evaluated. Primary immunodeficiency disorders comprise a group of more than one hundred diseases, several of which are fatal early in life. Yet, a majority of the patients are not diagnosed due to lack of high-throughput screening methods.

    METHODOLOGY/PRINCIPAL FINDINGS: We have previously developed a system using reverse phase protein microarrays for analysis of IgA levels in serum samples. In this study, we extended the applicability of the method to include determination of complement component C3 levels in eluates from DBSS collected at birth. Normal levels of C3 were readily detected in 269 DBSS from healthy newborns, while no C3 was detected in sera and DBSS from C3 deficient patients.

    CONCLUSIONS/SIGNIFICANCE: The findings suggest that patients with deficiencies of specific serum proteins can be identified by analysis of DBSS using reverse phase protein microarrays.

  • 10.
    Just, David
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science.
    Månberg, Anna
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics.
    Sjöberg, Ronald
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics.
    Burman, Joachim
    Nilsson, Peter
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics.
    Cunningham, Janet
    Exploring the autoantibody repertoire in patients with obsessive compulsive disorderManuscript (preprint) (Other academic)
  • 11.
    Qundos, Ulrika
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Drobin, Kimi
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mattsson, Cecilia
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hong, Mun-Gwan
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Forsström, Björn
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Solomon, David
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Uhlén, Mathias
    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.
    Michaelsson, Karl
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Affinity proteomics discovers decreased levels of AMFR in plasma from Osteoporosis patients2016In: PROTEOMICS - Clinical Applications, ISSN 1862-8346, E-ISSN 1862-8354, Vol. 10, no 6, p. 681-690Article in journal (Refereed)
    Abstract [en]

    Purpose: Affinity proteomic approaches by antibody bead arrays enable multiplexed analysis of proteins in body fluids. In the presented study, we investigated blood plasma within osteoporosis to discovery differential protein profiles and to propose novel biomarkers candidates for subsequent studies. Experimental design: Starting with 4608 antibodies and plasma samples from 22 women for an untargeted screening, a set of 72 proteins were suggested for further analysis. Complementing these with targets from literature and other studies, a targeted bead array of 180 antibodies was built to profile for 92 proteins in plasma samples of 180 women from two independent population-based studies. Results: Differential profiles between osteoporosis patients and matched controls were discovered for 12 proteins in at least one of the two study sets. Among these targets, the levels of autocrine motility factor receptor (AMFR) were concordantly lower in plasma of female osteoporosis patients. Subsequently, verification of anti-AMFR antibody selectivity was conducted using high-density peptide and protein arrays, and Western blotting. Conclusions and clinical relevance: Further validation in additional study sets will be needed to determine the clinical value of the observed decrease in AMFR plasma levels in osteoporosis patients, but AMFR may aid our understanding of disease mechanisms and could support existing tools for diagnosis and monitoring of patient mobility within osteoporosis.

  • 12.
    Rimini, Rebecca
    et al.
    KTH, School of Biotechnology (BIO), Proteomics.
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics.
    Sundberg, Marten
    Sjöberg, Ronald
    Klevebring, Daniel
    Gry, Marcus
    KTH, School of Biotechnology (BIO), Proteomics.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics.
    Validation of serum protein profiles by a dual antibody array approach2009Article in journal (Refereed)
    Abstract [en]

    In recent years, affinity-based technologies have become important tools for serum profiling to uncover protein expression patterns linked to disease state or therapeutic effects. In this study, we describe a path towards the production of an antibody microarray to allow protein profiling of biotinylated human serum samples with reproducible sensitivity in the picomolar range. With the availability of growing numbers of affinity reagents, protein profiles are to be validated in efficient manners and we describe a cross-platform strategy based on data concordance with a suspension bead array to interrogate the identical set of antibodies with the same cohort of serum samples. Comparative analysis enabled to screen for high-performing antibodies, which were displaying consistent results across the two platforms and targeting known serum components. Moreover, data processing methods such as sample referencing and normalization were evaluated for their effects on inter-platform agreement. Our work suggests that mutual validation of protein expression profiles using alternative microarray platforms holds great potential in becoming an important and valuable component in affinity-based high-throughput proteomic screenings as it allows to narrow down the number of discovered targets prior to orthogonal, uniplexed validation approaches.

  • 13. Sierra-Sanchez, Alvaro
    et al.
    Garrido-Martin, Diego
    Lourido, Lucia
    Gonzalez-Gonzalez, Maria
    Diez, Paula
    Ruiz-Romero, Cristina
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Droste, Conrad
    De Las Rivas, Javier
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Blanco, Francisco
    Fuentes, Manuel
    Screening and Validation of Novel Biomarkers in Osteoarticular Pathologies by Comprehensive Combination of Protein Array Technologies2017In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 16, no 5, p. 1890-1899Article in journal (Refereed)
    Abstract [en]

    Osteoarthritis (OA) is one of the most prevalent articular diseases. The identification of proteins closely associated with the diagnosis, progression, prognosis, and treatment response is dramatically required for this pathology. In this work, differential serum protein profiles have been identified in OA and rheumatoid arthritis (RA) by antibody arrays containing 151 antibodies against 121 antigens in a cohort of 36 samples. Then the identified differential serum protein profiles have been validated in a larger cohort of 282 samples. The overall immunoreactivity is higher in the pathological situations in comparison with the controls. Several proteins have been identified as biomarker candidates for OA and RA. Most of these biomarker candidates are proteins related to inflammatory response, lipid metabolism, or bone and extracellular matrix formation, degradation, or remodeling.

  • 14.
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    On Generation and Applications of High-Density Protein Microarrays2015Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Affinity proteomics has experienced rapid development over the last two decades and one of the most promising platforms to emerge are the protein microarrays. The combination of affinity reagents and miniaturisation enables assays for simultaneous high throughput and sensitive protein analysis. Due to the combination of these desrable properties, a multitude of protein array platforms for rapid and efficient study of proteomes and protein interactions are in use today. Although the protein microarray field has more than two decades of history to look back on the development of new protein microarray platforms continues to this day and beyond.

    In the paper I in this thesis, a microarray of eluates from dried blood spot samples collected from neonates were designed and utilised for detection of complement factor 3 (C3) deficiency. The data acquired from the microarrays platform were compared to C3 levels obtained through enzyme-linked immunosorbant assay (ELISA), and the microarray assay were found to separate the C3 deficient samples from the controls. The conclusion of this investigation was that the microarray platform would be suitable for high-throughput screening of C3 deficiency in neonates. Paper II outlines the work in developing a multiplex platform for validation of affinity reagents. A set of 398 affinity binders, originating from five research groups, were profiled against 432 antigens and representing both polyclonal rabbit antibodies, monoclonal mouse antibodies, and recombinant single-chain variable fragments. Approximately 50% of the binders were found to preferably recognise their intended target while 10% of the binders did not generate any, or low, signals with their respective targets. For paper III, a reverse phase array (RPPA) platform using fluorescence-based detection of IgA deficiency in over 2.000 samples where validated on a label-free detection system and ELISA. The data from the label-free platform and the RPPA were found agree well with each other while data from ELISA did with neither of them. It was found that the label-free platform proved to be well-suited for detection of IgA in serum. Paper IV describes one of the world’s largest protein microarrays containing 21.120 recombinant protein fragments. We describe some of the possible applications of these large-scale arrays, such as binding profiles for the validation of antibodies with 11.520 and 21.120 recombinant proteins, as well as screening for autoimmunity in human serum samples.

  • 15.
    Sjöberg, Ronald
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Andersson, Eni
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hellström, Cecilia
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mattsson, Cecilia
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Schwenk, Jochen M.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Nilsson, Peter
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Ayoglu, Burcu
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    High-density antigen microarrays for the assessment of antibody selectivity and off-target binding2018In: Epitope Mapping Protocols, Humana Press Inc. , 2018, p. 231-238Chapter in book (Refereed)
    Abstract [en]

    With the increasing availability of collections of antibodies, their evaluation in terms of binding selectivity becomes an important but challenging task. Planar antigen microarrays are very suitable tools to address this task and provide a powerful proteomics platform for the characterization of the binding selectivity of antibodies toward thousands of antigens in parallel. In this chapter, we describe our in-house developed procedures for the generation of high-density planar antigen microarrays with over 21,000 features. We also provide the details of the assay protocol, which we routinely use for the assessment of binding selectivity of the polyclonal antibodies generated within the Human Protein Atlas. © Springer Science+Business Media, LLC, part of Springer Nature 2018.

  • 16.
    Sjöberg, Ronald
    et al.
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hammarström, L.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Biosensor based protein profiling on reverse phase serum microarray2012In: Journal of Proteomics & Bioinformatics, ISSN 0974-276X, E-ISSN 0974-276X, Vol. 5, no 8, p. 185-189Article in journal (Refereed)
    Abstract [en]

    The reverse phase serum microarray format enables multi-parallel and simultaneous analysis of literally thousands of samples, a feature which is of uttermost importance for protein profiling of clinical samples. We have here screened 2400 serum samples for their potential IgA deficiency by using a fluorescence based reverse phase serum microarray platform and a biosensor based label-free microarray platform for verification and also compared our microarray-results to clinical routine ELISA. We have been able to identify possible IgA-deficiencies and to show the suitability of our microarray-platforms for large-scale screening of clinical serum samples. The two microarray methods show reproducibility and correlation towards each other and low variation between replicates within each platform. Both of the microarray platforms show less agreement towards ELISA. The fluorescence based microarray method has been shown to be applicable for large-scale screening of clinically important serum samples for detection of possibly IgA-deficient patients. Furthermore, it was found that the microarray based biosensor method could be used for determining the relative differences in concentration of IgA between the samples.

  • 17.
    Sjöberg, Ronald
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mattsson, Cecilia
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Andersson, Eni
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hellström, Cecilia
    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.
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Ayoglu, Burcu
    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.
    Exploration of high-density protein microarrays for antibody validation and autoimmunity profiling2016In: New Biotechnology, ISSN 1871-6784, E-ISSN 1876-4347, Vol. 33, no 5, p. 582-592Article in journal (Refereed)
    Abstract [en]

    High-density protein microarrays of recombinant human protein fragments, representing 12,412 unique Ensembl Gene IDs, have here been produced and explored. These protein microarrays were used to analyse antibody off-target interactions, as well as for profiling the human autoantibody repertoire in plasma against the antigens represented by the protein fragments. Affinity-purified polyclonal antibodies produced within the Human Protein Atlas (HPA) were analysed on microarrays of three different sizes, ranging from 384 antigens to 21,120 antigens, for evaluation of the antibody validation criteria in the HPA. Plasma samples from secondary progressive multiple sclerosis patients were also screened in order to explore the feasibility of these arrays for broad-scale profiling of autoantibody reactivity. Furthermore, analysis on these near proteome-wide microarrays was complemented with analysis on HuProt (TM) Human Proteome protein microarrays. The HPA recombinant protein microarray with 21,120 antigens and the HuProt (TM) Human Proteome protein microarray are currently the largest protein microarray platforms available to date. The results on these arrays show that the Human Protein Atlas antibodies have few off-target interactions if the antibody validation criteria are kept stringent and demonstrate that the HPA-produced high-density recombinant protein fragment microarrays allow for a high-throughput analysis of plasma for identification of possible autoantibody targets in the context of various autoimmune conditions.

  • 18.
    Sjöberg, Ronald
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Mattsson, Cecilia
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Andersson, Eni
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Hellström, Cecilia
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Zhu, Heng
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Ayoglu, Brucu
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Exploration of high-density protein microarrays for antibody validation and autoimmunity profilingManuscript (preprint) (Other academic)
  • 19.
    Sjöberg, Ronald
    et al.
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Sundberg, Mårten
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Gundberg, Anna
    KTH, School of Biotechnology (BIO).
    Sivertsson, Åsa
    KTH, School of Biotechnology (BIO).
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO).
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Validation of affinity reagents using antigen microarrays2011In: New Biotechnology, ISSN 1871-6784, E-ISSN 1876-4347, Vol. 29, no 5, p. 555-563Article in journal (Refereed)
    Abstract [en]

    There is a need for standardised validation of affinity reagents to determine their binding selectivity and specificity. This is of particular importance for systematic efforts that aim to cover the human proteome with different types of binding reagents. One such international program is the SH2-consortium, which was formed to generate a complete set of renewable affinity reagents to the SH2-domain containing human proteins. Here, we describe a microarray strategy to validate various affinity reagents, such as recombinant single-chain antibodies, mouse monoclonal antibodies and antigen-purified polyclonal antibodies using a highly multiplexed approach. An SH2-specific antigen microarray was designed and generated, containing more than 6000 spots displayed by 14 identical subarrays each with 406 antigens, where 105 of them represented SH2-domain containing proteins. Approximately 400 different affinity reagents of various types were analysed on these antigen microarrays carrying antigens of different types. The microarrays revealed not only very detailed specificity profiles for all the binders, but also showed that overlapping target sequences of spotted antigens were detected by off-target interactions. The presented study illustrates the feasibility of using antigen microarrays for integrative, high-throughput validation of various types of binders and antigens.

  • 20.
    Zhou, Xiamo
    et al.
    KTH, School of Electrical Engineering (EES), Micro and Nanosystems.
    Sjöberg, Ronald
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Druet, Amaury
    KTH, School of Electrical Engineering (EES), Micro and Nanosystems.
    Schwenk, Jochen M.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    van der Wijngaart, Wouter
    KTH, School of Electrical Engineering (EES), Micro and Nanosystems.
    Haraldsson, Tommy
    KTH, School of Electrical Engineering (EES), Micro and Nanosystems.
    Carlborg, Carl Fredrik
    KTH, School of Electrical Engineering (EES), Micro and Nanosystems.
    Thiol–ene–epoxy thermoset for low-temperature bonding to biofunctionalized microarray surfaces2017In: Lab on a Chip, ISSN 1473-0197, E-ISSN 1473-0189, Vol. 17, no 21, p. 3672-3681Article in journal (Refereed)
    Abstract [en]

    One way to improve the sensitivity and throughput of miniaturized biomolecular assays is to integrate microfluidics to enhance the transport efficiency of biomolecules to the reaction sites. Such microfluidic integration requires bonding of a prefabricated microfluidic gasket to an assay surface without destroying its biological activity. In this paper we address the largely unmet challenge to accomplish a proper seal between a microfluidic gasket and a protein surface, with maintained biological activity and without contaminating the surface or blocking the microfluidic channels. We introduce a novel dual cure polymer resin for the formation of microfluidic gaskets that can be room-temperature bonded to a range of substrates using only UVA light. This polymer is the first polymer that features over a month of shelf life between the structure formation and the bonding, moreover the fully cured polymer gaskets feature the following set of properties suitable for microfluidics: high stiffness, which prevents microfluidic channel collapse during handling; very limited absorption of biomolecules; and no significant leaching of uncured monomers. We describe the novel polymer resin and its characteristics, study through FT-IR, and demonstrate its use as microfluidic well-arrays bonded onto protein array slides at room temperature followed by multiplexed immunoassays. The results confirm maintained biological activity and show high repeatability between protein arrays. This new approach for integrating microfluidic gaskets to biofunctionalised surfaces has the potential to improve sample throughput and decrease manufacturing costs for miniaturized biomolecular systems.

1 - 20 of 20
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