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Bead-based and multiplexed immunoassays for protein profiling via sequential affinity capture
KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
2017 (English)In: Serum/Plasma Proteomics, Humana Press, 2017, p. 45-54Chapter in book (Refereed)
Abstract [en]

Antibody microarrays offer high-throughput immunoassays for multiplexed analyses of clinical samples. For such approaches, samples are either labeled in solution to enable a direct readout on the single binder assay format or detected by matched pairs of capture and detection antibodies in dual binder assay format, also known as sandwich assays. Aiming to benefit from the flexibility and capacity offered by single binder assay readout and the specificity and sensitivity of dual binder assays, we developed a multiplexed dual binder procedure that is based on a sequential, rather than combined, antigen binding. The method, entitled dual capture assay (DCA), is composed of an initial antigen capture by antibodies on beads, followed by labeling of captured protein targets on beads, combinatorial elution steps at high and low pH, and a readout using a secondary bead array. Compared to classical single binder assays, the described method demonstrated several advantages such as reduced contribution of off-target binding, lower noise levels, and improved correlation when comparing with clinical reference values. This procedure describes a novel and versatile immunoassay strategy for proteome profiling in body fluids.

Place, publisher, year, edition, pages
Humana Press, 2017. p. 45-54
Series
Methods in Molecular Biology, ISSN 1064-3745 ; 1619
Keywords [en]
Affinity proteomics, Antibody arrays, Antibody selectivity, Assay sensitivity, Plasma profiling, Suspension bead array
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:kth:diva-211884DOI: 10.1007/978-1-4939-7057-5_4Scopus ID: 2-s2.0-85021745527ISBN: 9781493970568 (print)OAI: oai:DiVA.org:kth-211884DiVA, id: diva2:1131634
Funder
Knut and Alice Wallenberg FoundationScience for Life Laboratory - a national resource center for high-throughput molecular bioscience
Note

QC 20170815

Available from: 2017-08-15 Created: 2017-08-15 Last updated: 2017-08-15Bibliographically approved

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