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Dissecting Antibodies with Regards to Linear and Conformational Epitopes
KTH, Centres, Science for Life Laboratory, SciLifeLab.
KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
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2015 (English)In: PLoS ONE, ISSN 1932-6203, Vol. 10, no 3, e0121673Article in journal (Refereed) Published
Abstract [en]

An important issue for the performance and specificity of an antibody is the nature of the binding to its protein target, including if the recognition involves linear or conformational epitopes. Here, we dissect polyclonal sera by creating epitope-specific antibody fractions using a combination of epitope mapping and an affinity capture approach involving both synthesized peptides and recombinant protein fragments. This allowed us to study the relative amounts of antibodies to linear and conformational epitopes in the polyclonal sera as well as the ability of each antibody-fraction to detect its target protein in Western blot assays. The majority of the analyzed polyclonal sera were found to have most of the target-specific antibodies directed towards linear epitopes and these were in many cases giving Western blot bands of correct molecular weight. In contrast, many of the antibodies towards conformational epitopes did not bind their target proteins in the Western blot assays. The results from this work have given us insights regarding the nature of the antibody response generated by immunization with recombinant protein fragments and has demonstrated the advantage of using antibodies recognizing linear epitopes for immunoassay involving wholly or partially denatured protein targets.

Place, publisher, year, edition, pages
2015. Vol. 10, no 3, e0121673
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Medical Biotechnology
URN: urn:nbn:se:kth:diva-166504DOI: 10.1371/journal.pone.0121673ISI: 000352133600110PubMedID: 25816293ScopusID: 2-s2.0-84926388037OAI: diva2:811817
Knut and Alice Wallenberg Foundation

QC 20150513

Available from: 2015-05-13 Created: 2015-05-11 Last updated: 2015-05-13Bibliographically approved

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Forsström, BjörnAxnäs, Barbara BislawskaRockberg, JohanDanielsson, HannaBohlin, AnnaUhlén, Mathias
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