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Seed amplification assays for diagnosing synucleinopathies: the issue of influencing factors

Authors :
Giovanni Bellomo
Silvia Paciotti
Leonardo Gatticchi
Domenico Rizzo
Federico Paolini Paoletti
Marco Fragai
Lucilla Parnetti
Source :
Frontiers in Bioscience-Landmark, Vol 26, Iss 11, Pp 1075-1088 (2021)
Publication Year :
2021
Publisher :
IMR Press, 2021.

Abstract

Background: The prion-like misfolding and aggregation of α-synuclein (α-syn) is involved in the pathophysiology of Parkinson’s disease and other synucleinopathies. Seed amplification assays (SAAs) are biophysical tools that take advantage on the peculiar properties of prion proteins by amplifying small amounts of aggregates in biological fluids at the expense of recombinant monomeric protein added in solution. SAAs have emerged as the most promising tools for the diagnosis of synucleinopathies in vivo. However, the diagnostic outcome of SAAs depends on the aggregation kinetics of α-syn, which in turn is influenced by several experimental variables. Methods: In our work, we analysed the impact on SAAs of some of the most critical experimental factors by considering models that describe the aggregation kinetics of α-syn. Results: We started our analysis by making simulations to understand which kinetic models could explain the aggregation kinetics of α-syn during incubation/shaking cycles. Subsequently, under shaking/incubation cycles similar to the ones commonly used in SAAs, we tested the influence of some analytical variables such as monomer concentration, presence/absence of glass beads, pH, addition of human cerebrospinal fluid, and use of detergents on α-syn aggregation. Conclusions: Our investigation highlighted how optimization and standardization of experimental procedures for α-syn SAAs is of utmost relevance for the ultimate goal of applying these assays in clinical routine. Although these aspects have been evaluated with specific SAA protocols, most of the experimental variables considered influenced very general aggregation mechanisms of α-syn, thus making most of the results obtained from our analyses extendable to other protocols.

Details

Language :
English
ISSN :
27686701
Volume :
26
Issue :
11
Database :
Directory of Open Access Journals
Journal :
Frontiers in Bioscience-Landmark
Publication Type :
Academic Journal
Accession number :
edsdoj.36a8e4382db4a64a4584b07b425bc20
Document Type :
article
Full Text :
https://doi.org/10.52586/5010