Biofunctionalized Silica Nanoparticles: Standards in Amyloid-β Oligomer-Based Diagnosis of Alzheimer’s Disease

Abstract

Amyloid-β (Aβ) oligomers represent a promising biomarker for the early diagnosis of Alzheimer's disease (AD). However, state-of-the-art methods for immunodetection of Aβ oligomers in body fluids show a large variability and lack a reliable and stable standard that enables the reproducible quantitation of Aβ oligomers. At present, the only available standard applied in these assays is based on a random aggregation process of synthetic Aβ and has neither a defined size nor a known number of epitopes. In this report, we generated a highly stable standard in the size range of native Aβ oligomers that exposes a defined number of epitopes. The standard consists of a silica nanoparticle (SiNaP), which is functionalized with Aβ peptides on its surface (Aβ-SiNaP). The different steps of Aβ-SiNaP synthesis were followed by microscopic, spectroscopic and biochemical analyses. To investigate the performance of Aβ-SiNaPs as an appropriate standard in Aβ oligomer immunodetection, Aβ-SiNaPs were diluted in cerebrospinal fluid and quantified down to a concentration of 10 fM in the sFIDA (surface-based fluorescence intensity distribution analysis) assay. This detection limit corresponds to an Aβ concentration of 1.9 ng l-1 and lies in the sensitivity range of currently applied diagnostic tools based on Aβ oligomer quantitation. Thus, we developed a highly stable and well-characterized standard for the application in Aβ oligomer immunodetection assays that finally allows the reproducible quantitation of Aβ oligomers down to single molecule level and provides a fundamental improvement for the worldwide standardization process of diagnostic methods in AD research.

Advancements of the sFIDA method for oligomer‐based diagnostics of neurodegenerative diseases

“…Since then, the sFIDA method underwent fundamental optimization in respect to automation , adaptation to blood‐based analysis and development of a suitable calibration standards .…”

“…A second route for obtaining stable standards is to use bioconjugated nanoparticles . In this method, nanoparticles with a narrow size range are synthesized as core material, which determines the final size of the oligomer (Fig.…”

“…The limitations of the method are caused by the positions of primary amine groups in the protein monomer which are bound to the particle surface. Ideally, the amines lie within the aggregation region of the protein, which is the case for Aβ and α‐synuclein . In such cases, the aggregation of the protein monomers on the particle surface is hampered.…”

Analysis of anticoagulants for blood-based quantitation of amyloid β oligomers in the sFIDA assay

“…Aβ-SiNaPs were prepared as described previously (Hülsemann et al, 2016). Briefly, bare silica nanoparticles (SiNaPs) were synthesized via Stöber process and silanized with APTES to cover the surface with primary amino groups.…”

“…Aβ-SiNaPs have been shown to be a suitable standard in Aβ oligomer quantitation due to their uniform size and defined number of accessible epitopes. Moreover, the particles are not prone to aggregation and are stable over several months (Hülsemann et al, 2016).…”