Separation and characterization of aggregated species of amyloid-beta peptides
2010 (English)In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 397, no 6, 2357-2366 p.Article in journal (Refereed) Published
We have investigated the use of isoelectric focusing and immunodetection for the separation of low molecular weight species of amyloid-beta (A beta) peptides from their aggregates. From solutions of A beta(1-40) or A beta(1-42) monomeric peptides, low molecular weight material appeared at a pI value of ca. 5, while the presence of aggregates was detected as bands, observed at a pI of 6-6.5. The formation of A beta aggregates (protofibrils) was verified by a sandwich ELISA, employing the protofibril conformation-selective antibody mAb158. In order to study the aggregation behavior when using a mixture of the monomers, we utilized the IEF separation combined with Western blot using two polyclonal antisera, selective for A beta(1-40) and A beta(1-42), respectively. We conclude that both monomers were incorporated in the aggregates. In a further study of the mixed aggregates, we used the protofibril conformation-selective antibody mAb158 for immunoprecipitation, followed by nanoelectrospray mass spectrometry (IP-MS). This showed that the A beta(1-42) peptide is incorporated in the aggregate in a significantly larger proportion than its relative presence in the original monomer composition. IP-MS with mAb158 was also performed, and compared to IP-MS with the A beta-selective antibody mAb1C3, where a monomeric A beta(1-16) peptide was added to the protofibril preparation. A beta(1-16) is known for its poor aggregation propensity, and acted therefore as a selectivity marker. The results obtained confirmed the protofibril conformation selectivity of mAb158.
Place, publisher, year, edition, pages
2010. Vol. 397, no 6, 2357-2366 p.
Alzheimer's disease, Protofibrils, Amyloid-beta (A beta), Isoelectric focusing (IEF)., Mass spectrometry (MS), Bioanalytical methods, Capillary electrophoresis / Electrophoresis
IdentifiersURN: urn:nbn:se:kth:diva-26073DOI: 10.1007/s00216-010-3839-9ISI: 000279453000038ScopusID: 2-s2.0-77955981824OAI: oai:DiVA.org:kth-26073DiVA: diva2:369695
6th International Conference on Instrumental Methods of Analysis Athens, GREECE, OCT 04-08, 2009
QC 201102142010-11-112010-11-112011-06-15Bibliographically approved