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Cytokine detection by antibody-based proximity ligation
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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2004 (English)In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 101, no 22, 8420-4 p.Article in journal (Other academic) Published
Abstract [en]

Efficient and precise detection techniques, along with extensive repertoires of specific binding reagents, will be needed to meet the challenges of proteome analyses. The recently established proximity ligation mechanism enables sensitive high-capacity protein measurements by converting the detection of specific proteins to the analysis of DNA sequences. Proximity probes containing oligonucleotide extensions are designed to bind pairwise to target proteins and to form amplifiable tag sequences by ligation when brought in proximity. In our previous report, both the ligatable arms and the protein binders were DNA molecules. We now generalize the method by providing simple and convenient protocols to convert any polyclonal antibodies or matched pair of monoclonal antibodies to proximity probe sets through the attachment of oligonucleotide sequences. Sufficient reagent for >100,000 proximity ligation assays can be prepared from 1 microg of antibody. The technique is applied to measure cytokines in a homogenous test format with femtomolar detection sensitivities in 1-microl samples, and we exemplify its utility in situations when only minute sample amounts are available.

Place, publisher, year, edition, pages
2004. Vol. 101, no 22, 8420-4 p.
Keyword [en]
Antibodies/*metabolism, Cell Line, Comparative Study, Cytokines/*analysis, Humans, Insulin/analysis, Molecular Probe Techniques, Oligonucleotides/metabolism, Research Support; Non-U.S. Gov't, Thrombin/analysis
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-70638DOI: 10.1073/pnas.0400552101PubMedID: 15155907OAI: oai:DiVA.org:uu-70638DiVA: diva2:98549
Available from: 2005-04-26 Created: 2005-04-26 Last updated: 2017-11-21Bibliographically approved

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Gullberg, MatsSchallmeiner, EdithJarvius, JonasLandegren, Ulf

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