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Analysis of protein expression in cell microarrays: A tool for antibody-based proteomics
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
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2006 (English)In: Journal of Histochemistry and Cytochemistry, ISSN 0022-1554, E-ISSN 1551-5044, Vol. 54, no 12, 1413-1423 p.Article in journal (Refereed) Published
Abstract [en]

Tissue microarray (TMA) technology provides a possibility to explore protein expression patterns in a multitude of normal and disease tissues in a high-throughput setting. Although TMAs have been used for analysis of tissue samples, robust methods for studying in vitro cultured cell lines and cell aspirates in a TMA format have been lacking. We have adopted a technique to homogeneously distribute cells in an agarose gel matrix, creating an artificial tissue. This enables simultaneous profiling of protein expression in suspension- and adherent-grown cell samples assembled in a microarray. In addition, the present study provides an optimized strategy for the basic laboratory steps to efficiently produce TMAs. Presented modifications resulted in an improved quality of specimens and a higher section yield compared with standard TMA production protocols. Sections from the generated cell TMAs were tested for immunohistochemical staining properties using 20 well-characterized antibodies. Comparison of immunoreactivity in cultured dispersed cells and corresponding cells in tissue samples showed congruent results for all tested antibodies. We conclude that a modified TIVIA technique, including cell samples, provides a valuable tool for high-throughput analysis of protein expression, and that this technique can be used for global approaches to explore the human proteome.

Place, publisher, year, edition, pages
2006. Vol. 54, no 12, 1413-1423 p.
Keyword [en]
immunohistochemistry, cell line, tissue microarray, affinity proteomics, antibody-based proteomics
National Category
Medical and Health Sciences
URN: urn:nbn:se:uu:diva-97142DOI: 10.1369/jhc.6A7001.2006ISI: 000242056300010PubMedID: 16957166OAI: oai:DiVA.org:uu-97142DiVA: diva2:171950
Available from: 2008-04-25 Created: 2008-04-25 Last updated: 2011-05-12Bibliographically approved
In thesis
1. Antibody-based Profiling of Expression Patterns using Cell and Tissue Microarrays
Open this publication in new window or tab >>Antibody-based Profiling of Expression Patterns using Cell and Tissue Microarrays
2008 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

In this thesis, methods to study gene and protein expression in cells and tissues were developed and utilized in combination with protein-specific antibodies, with the overall objective to attain greater understanding of protein function.

To analyze protein expression in in vitro cultured cell lines, a cell microarray (CMA) was developed, facilitating antibody-based protein profiling of cell lines using immunohistochemistry (IHC). Staining patterns in cell lines were analyzed using image analysis, developed to automatically identify cells and immunohistochemical staining, providing qualitative and quantitative measurements of protein expression. Quantitative IHC data from CMAs stained with nearly 3000 antibodies was used to evaluate the adequacy of using cell lines as models for cancer tissue. We found that cell lines are homogenous with respect to protein expression profiles, and generally more alike each other, than corresponding cancer cells in vivo. However, we found variability between cell lines in regards to the level of retained tumor phenotypic traits, and identified cell lines with a preserved link to corresponding cancer, suggesting that some cell lines are appropriate model systems for specific tumor types.

Specific gene expression patterns were analyzed in vitiligo vulgaris and malignant melanoma. Transcriptional profiling of vitiligo melanocytes revealed dysregulation of genes involved in melanin biosynthesis and melanosome function, thus highlighting some mechanisms possibly involved in the pathogenesis of vitiligo. Two new potential markers for infiltrating malignant melanoma, Syntaxin-7 and Discs large homolog 5, were identified using antibody-based protein profiling of melanoma in a tissue microarray format. Both proteins were expressed with high specificity in melanocytic lesions, and loss of Syntaxin-7 expression was associated with more high-grade malignant melanomas.

In conclusion, the combination of antibody-based proteomics and microarray technology provided valuable information of expression patterns in cells and tissues, which can be used to better understand associations between protein signatures and disease.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2008. 51 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 344
Pathology, antibody-based proteomics, tissue microarray, cell line, immunohistochemistry, melanocytes, image analysis, biomarker, Patologi
urn:nbn:se:uu:diva-8680 (URN)978-91-554-7183-5 (ISBN)
Public defence
2008-05-16, Rudbecksalen, Rudbecklaboratoriet, Dag Hammarskjöldsväg 20, Uppsala, 10:00
Available from: 2008-04-25 Created: 2008-04-25Bibliographically approved

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Strömberg, SaraKampf, CarolineWester, KennethPontén, Fredrik
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