1. |
- Berglund, Lisa, et al.
(författare)
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A genecentric Human Protein Atlas for expression profiles based on antibodies
- 2008
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Ingår i: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 7:10, s. 2019-2027
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Forskningsöversikt (refereegranskat)abstract
- 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 approximately 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.
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2. |
- Pontén, Fredrik, et al.
(författare)
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A global view of protein expression in human cells, tissues, and organs
- 2009
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Ingår i: Molecular Systems Biology. - : EMBO. - 1744-4292 .- 1744-4292. ; 5
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Tidskriftsartikel (refereegranskat)abstract
- 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
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3. |
- Uhlén, Mathias, et al.
(författare)
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A human protein atlas for normal and cancer tissues based on antibody proteomics
- 2005
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Ingår i: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 4:12, s. 1920-1932
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Tidskriftsartikel (refereegranskat)abstract
- 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.
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4. |
- Uhlén, Mathias, et al.
(författare)
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Towards a knowledge-based Human Protein Atlas
- 2010
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Ingår i: Nature Biotechnology. - : Springer Science and Business Media LLC. - 1087-0156 .- 1546-1696. ; 28:12, s. 1248-1250
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Tidskriftsartikel (refereegranskat)
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5. |
- Wester, Kenneth, et al.
(författare)
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Zinc-based fixative improves preservation of genomic DNA and proteins in histoprocessing of human tissues.
- 2003
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Ingår i: Lab Invest. - 0023-6837. ; 83:6, s. 889-99
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Tidskriftsartikel (refereegranskat)abstract
- Advantageous preservation of histology and detailed cellular morphology has rendered neutral buffered formalin (NBF) the most widely used fixative in clinical pathology. Despite excellent morphology for routine diagnostics, a major drawback of NBF fixation is its detrimental effect on DNA and RNA quality. In addition to complicating analysis of genes and transcripts in complex tissues, NBF denatures proteins and thereby hampers immunohistochemical visualization of certain antigens. In the present study, we evaluated a zinc-based fixative (ZBF) regarding its effects on tissue morphology, quality of genomic DNA, and preservation of protein immunoreactivity in a broad spectrum of tissues. Four different modes of fixation were analyzed: ZBF-paraffin embedding, NBF-paraffin embedding, ZBF-fixation prior to snap-freezing, and immediate snap-freezing. Laser-assisted microdissection, allowing retrieval of a defined number of cells for PCR, was used to study DNA quality. Genomic DNA was analyzed using primers for beta2-microglobulin and the transferrin receptor. Immunohistochemistry was performed using nine antibodies. Tissue microarray blocks were used for analysis of morphology and immunoreactivity. Only slight impairment of morphologic qualities was found after ZBF-paraffin embedding, whereas ZBF prior to freezing resulted in a more crisp morphology compared with routine cryosections. A significantly higher DNA yield was observed in samples isolated from ZBF-paraffin-embedded tissues compared with NBF-paraffin-embedded tissues. Both yield and quality of DNA was comparable in frozen tissues irrespective to prior ZBF fixation. Immunoreactivity in paraffin-embedded tissue was superior in ZBF-fixated tissue compared with NBF-fixated for a majority of tested antibodies. Furthermore, for seven out of nine antibodies, antigen retrieval pretreatment proved unnecessary in ZBF-fixated tissue. Thus, despite a slight impairment of morphology, ZBF preserves protein structures well. We conclude that ZBF is superior to NBF for analysis of DNA and protein expression. Fixation of tissues in ZBF may also be an alternative strategy to freeze storage of tissue specimens, eg, in future bio-banks.
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