title: Luminometric Nanoparticle-Based Assay for High Sensitivity Detection of β-Amyloid Aggregation.
abstract: A nanoparticle-based assay utilizing time-resolved luminescence resonance energy transfer (TR-LRET) was developed for the detection of β-amyloid aggregation. The assay is based on the competitive adsorption of the sample and the acceptor-labeled protein to donor europium(III) polystyrene nanoparticles. The performance of the assay was demonstrated by following the fibrillization of β-amyloid peptide 1-42 (Aβ42) as a function of time and by comparing to the reference methods atomic force microscopy (AFM) and thioflavin T (ThT) assay. The fibrillization leads to reduced adsorption of Aβ42to the nanoparticles increasing the TR-LRET signal. The investigated methods detected fibril formation with equal sensitivities. Eight potential fibrillization inhibitor compounds reported in the literature were tested and the results obtained with each method were compared. It was shown with AFM imaging that the inhibition of fibril formation was not complete with any of the compounds. The developed TR-LRET nanoparticle assay gave corresponding results with the AFM imaging. However, the ThT assay led to contradictory results, as low fluorescence signal was measured in the presence of all tested compounds suggesting inhibition of fibrillization. Our results suggest that the developed TR-LRET nanoparticle assay can be exploited for screening of potential β-amyloid aggregation inhibitors, whereas some of the tested compounds may be measured as false positive inhibitors with the much-utilized ThT assay.
authors.affiliation: Laboratory of Materials Chemistry and Chemical Analysis, Department of Chemistry, University of Turku , Vatselankatu 2, FI-20500 Turku, Finland.|Laboratory of Biophysics, Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Tykistökatu 6A, FI-20520 Turku, Finland.
authors.forename: Sari
authors.identifier: 0000-0002-2411-6724
authors.initials: S
authors.lastname: Pihlasalo
authors.affiliation: Laboratory of Biophysics, Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Tykistökatu 6A, FI-20520 Turku, Finland.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Deguchi
authors.affiliation: Laboratory of Biophysics, Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Tykistökatu 6A, FI-20520 Turku, Finland.
authors.forename: Maria
authors.initials: M
authors.lastname: Virtamo
authors.affiliation: Laboratory of Biophysics, Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Tykistökatu 6A, FI-20520 Turku, Finland.
authors.forename: Jenna
authors.initials: J
authors.lastname: Jacobino
authors.affiliation: Laboratory of Biophysics, Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Tykistökatu 6A, FI-20520 Turku, Finland.
authors.forename: Karthik
authors.initials: K
authors.lastname: Chary
authors.affiliation: PET Preclinical Imaging Laboratory, Turku PET Centre, University of Turku , Itäinen Pitkäkatu 4B, FI-20520 Turku, Finland.
authors.forename: Francisco R
authors.initials: FR
authors.lastname: López-Picón
authors.affiliation: Department of Medicinal Chemistry, University of Vienna , Althanstrasse 14, A-1090 Vienna, Austria.
authors.forename: Gerda
authors.initials: G
authors.lastname: Brunhofer-Bolzer
authors.affiliation: Laboratory of Biophysics, Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Tykistökatu 6A, FI-20520 Turku, Finland.
authors.forename: Roope
authors.initials: R
authors.lastname: Huttunen
authors.affiliation: Pharmaceutical Design and Discovery Group (PharmDD), Division of Pharmaceutical Biosciences, Faculty of Pharmacy, University of Helsinki , Viikinkaari 5E, FI-00014 Helsinki, Finland.
authors.forename: Adyary
authors.initials: A
authors.lastname: Fallarero
authors.affiliation: Pharmaceutical Design and Discovery Group (PharmDD), Division of Pharmaceutical Biosciences, Faculty of Pharmacy, University of Helsinki , Viikinkaari 5E, FI-00014 Helsinki, Finland.
authors.forename: Pia
authors.initials: P
authors.lastname: Vuorela
authors.affiliation: Laboratory of Materials Chemistry and Chemical Analysis, Department of Chemistry, University of Turku , Vatselankatu 2, FI-20500 Turku, Finland.|Laboratory of Biophysics, Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Tykistökatu 6A, FI-20520 Turku, Finland.
authors.forename: Harri
authors.initials: H
authors.lastname: Härmä
mesh_terms: D016229:Amyloid beta-Peptides / Q000032:analysis* / Q000037:antagonists & inhibitors / Q000378:metabolism; D053119:Benzophenanthridines / Q000737:chemistry / Q000378:metabolism; D005063:Europium / Q000737:chemistry; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry; D018625:Microscopy, Atomic Force; D053758:Nanoparticles / Q000737:chemistry* / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis* / Q000037:antagonists & inhibitors / Q000378:metabolism; D011137:Polystyrenes / Q000737:chemistry; D066329:Protein Aggregates; D012293:Rifampin / Q000737:chemistry / Q000378:metabolism
chemical_list: D016229:Amyloid beta-Peptides; D053119:Benzophenanthridines; D005456:Fluorescent Dyes; D010446:Peptide Fragments; D011137:Polystyrenes; D066329:Protein Aggregates; C075222:amyloid beta-protein (1-42); D005063:Europium; C016299:chelerythrine; D012293:Rifampin