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| 2. |
- Andersson, Sandra, et al.
(author)
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Insufficient antibody validation challenges oestrogen receptor beta research
- 2017
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In: Nature Communications. - : NATURE PUBLISHING GROUP. - 2041-1723. ; 8
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Journal article (peer-reviewed)abstract
- The discovery of oestrogen receptor beta (ER beta/ESR2) was a landmark discovery. Its reported expression and homology with breast cancer pharmacological target ER alpha (ESR1) raised hopes for improved endocrine therapies. After 20 years of intense research, this has not materialized. We here perform a rigorous validation of 13 anti-ER beta antibodies, using well-characterized controls and a panel of validation methods. We conclude that only one antibody, the rarely used monoclonal PPZ0506, specifically targets ER beta in immunohistochemistry. Applying this antibody for protein expression profiling in 44 normal and 21 malignant human tissues, we detect ER beta protein in testis, ovary, lymphoid cells, granulosa cell tumours, and a subset of malignant melanoma and thyroid cancers. We do not find evidence of expression in normal or cancerous human breast. This expression pattern aligns well with RNA-seq data, but contradicts a multitude of studies. Our study highlights how inadequately validated antibodies can lead an exciting field astray.
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| 4. |
- Colwill, Karen, et al.
(author)
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A roadmap to generate renewable protein binders to the human proteome
- 2011
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In: Nature Methods. - : Nature America Inc.. - 1548-7091 .- 1548-7105. ; 8:7, s. 551-8
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Journal article (peer-reviewed)abstract
- 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.
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| 5. |
- Eriksson, Ola, et al.
(author)
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Enhancement of biogas production from food waste and sewage sludge : environmental and economic life cycle performance
- 2016
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In: Journal of Environmental Management. - : Elsevier BV. - 0301-4797 .- 1095-8630. ; 175, s. 33-39
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Journal article (peer-reviewed)abstract
- Management of municipal solid waste is an efficient method to increase resource efficiency, as well as to replace fossil fuels with renewable energy sources due to that (1) waste to a large extent is renewable as it consists of food waste, paper, wood etc. and (2) when energy and materials are recovered from waste treatment, fossil fuels can be substituted. In this paper results from a comprehensive system study of future biological treatment of readily degradable waste in two Swedish regions are presented. Different collection and separation systems for food waste in households have been applied as well as technical improvements of the biogas process as to reduce environmental impact. The results show that central sorting of a mixed fraction into recyclables, combustibles, biowaste and inert is a competitive option compared to source separation. Use of pellets is beneficial compared to direct spreading as fertiliser. Fuel pellets seem to be the most favourable option, which to a large extent depends on the circumstances in the energy system. Separation and utilisation of nitrogen in the wet part of the digestion residue is made possible with a number of technologies which decreases environmental impact drastically, however to a substantial cost in some cases.
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| 6. |
- Eriksson, Ola, et al.
(author)
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Integrated waste management as a mean to promote renewable energy
- 2014
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In: Renewable energy. - : Elsevier BV. - 0960-1481 .- 1879-0682. ; 61, s. 38-42
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Journal article (peer-reviewed)abstract
- Management of municipal solid waste is an efficient method to both increase resource efficiency (material and energy recovery instead of landfill disposal) and to replace fossil fuels with renewable energy sources (waste is renewable in itself to a large extent as it contains paper, wood, food waste etc.). The paper presents the general outline and results from a comprehensive system study of future waste management. In the study a multifunctional waste management system integrated with local energy systems for district heating and electricity, wastewater treatment, agriculture and vehicle fuel production is investigated with respect to environmental impact and financial economy. Different waste technologies as well as management strategies have been tested. The treatment is facilitated through advanced sorting, efficient treatment facilities and upgrading of output products. Tools used are the ORWARE model for the waste management system and the MARTES model for the district heating system. The results for potential global warming are used as an indicator for renewable energy. In all future scenarios and for all management strategies net savings of CO2 is accomplished. Compared to a future reference the financial costs will be higher or lower depending on management strategy.
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| 7. |
- Gantelius, Jesper, et al.
(author)
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A ten-minute high density lateral flow protein microarray assay
- 2011
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In: 15th International Conference on Miniaturized Systems for Chemistry and Life Sciences 2011. - 9781618395955 ; , s. 1176-1178
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Conference paper (peer-reviewed)abstract
- 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.
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| 8. |
- Janzi, M., et al.
(author)
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Serum microarrays for large scale screening of protein levels
- 2005
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In: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 4:12, s. 1942-1947
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Journal article (peer-reviewed)abstract
- There is a great need for comprehensive proteomic analysis of large patient cohorts of plasma and serum samples to identify biomarkers of human diseases. Here we describe a new antibody-based proteomic approach involving a reverse array format where serum samples are spotted on a microarray. This enables all samples to be screened for their content of a certain serum protein in a single experiment using target-recognizing antibodies and fluorescently labeled secondary antibodies. The procedure is illustrated with the analysis of the IgA levels in 2009 spotted serum samples, and the data are compared with clinical routine measurements. The results suggest that it is possible to simultaneously screen thousands of complex clinical serum samples for their content of the relative amount of specific serum proteins of clinical relevance.
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| 9. |
- Katona, Borbala, et al.
(author)
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Antibody Validation Strategy for Nuclear Receptors.
- 2019
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In: Methods in Molecular Biology. - New York, NY : Springer New York. - 1064-3745 .- 1940-6029. ; 1966, s. 79-99
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Journal article (peer-reviewed)abstract
- Antibodies are invaluable biological tools that we can use to detect the presence, location, or alteration of nuclear receptors. However, antibodies frequently cross-react with other proteins and their performance can vary from batch to batch, from application to application and from lab to lab. When each lot of antibody is not thoroughly validated for each assay, each sample type, and each lab and user, antibody-based assays can lead to flawed interpretations and reproducibility problems. In this chapter, we describe a scheme for thorough antibody validation, suitable for nuclear receptors. The method is based on using highly characterized positive and negative controls assembled into a validation tissue microarray (TMA). Through correlation of immunohistochemical staining (IHC) and mRNA levels over multiple tissues, use of current public databases, and assessment of binding to intended and nonintended targets using western blotting (WB), immunoprecipitation (IP), and mass spectrometry (MS), we describe a path for thoroughly validation of antibodies.
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| 10. |
- Larsson, Mårten, et al.
(author)
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Selective interaction of megalin with postsynaptic density-95 (PSD-95)-like membrane-associated guanylate kinase (MAGUK) proteins
- 2003
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In: Biochemical Journal. - 0264-6021 .- 1470-8728. ; 373:2, s. 381-391
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Journal article (peer-reviewed)abstract
- Megalin is an integral membrane receptor belonging to the low-density lipoprotein receptor family. In addition to its role as an endocytotic receptor, megalin has also been proposed to have signalling functions. Using interaction cloning in yeast, we identified the membrane-associated guanylate kinase family member postsynaptic density-95 (PSD-95) as an interaction partner for megalin. PSD-95 and a truncated version of megalin were co-immunoprecipitated from HEK-293 cell lysates overexpressing the two proteins, which confirmed the interaction. The two proteins were found to be co-localized in these cells by confocal microscopy. Immunocytochemical studies showed that cells in the parathyroid, proximal tubuli of the kidney and placenta express both megalin and PSD-95. We found that the interaction between the two proteins is mediated by the binding of the C-terminus of megalin, which has a type I PSD-95/ Drosophila discs-large/zona occludens 1 (PDZ)-binding motif, to the PDZ2 domain of PSD-95. The PSD-95-like membrane-associated guanylate kinase ('MAGUK') family contains three additional members: PSD-93, synapse-associated protein 97 (SAP97) and SAP102. We detected these proteins, apart from SAP102, in parathyroid chief cells, a cell type having a marked expression of megalin. The PDZ2 domains of PSD-93 and SAP102 were also shown to interact with megalin, whereas no interaction was detected for SAP97. The SAP97 PDZ2 domain differed at four positions from the other members of the PSD-95 subfamily. One of these residues was Thr(389), located in the alphaB-helix and part of the hydrophobic pocket of the PDZ2 domain. Surface plasmon resonance experiments revealed that mutation of SAP97 Thr(389) to alanine, as with the other PSD-95-like membrane-associated guanylate kinases, induced binding to megalin.
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