Ultrasensitive Detection of Aβ42 Seeds in Cerebrospinal Fluid with a Nanopipette-Based Real-Time Fast Amyloid Seeding and Translocation Assay
Résumé
In this work, early-stage Aβ42 aggregates were detected using a real-time Fast Amyloid Seeding and Translocation (RT-FAST) assay. Specifically, Aβ42 monomers were incubated in buffer solution with and without preformed Aβ42 seeds in a quartz nanopipette coated with L-DOPA. Then, formed Aβ42 aggregates were analysed on flyby resistive pulse sensing at various incubation time points. Aβ42 aggregates were detected only in the sample with Aβ42 seeds after 180 minutes of incubation, giving an on/off readout of the presence of preformed seeds. Moreover, this RT-FAST assay could detect preformed seeds spiked in 4% cerebrospinal fluid/buffer solution. However, in this condition, the time to detect the aggregates and the nanopipette lifetime were increased. Analysis of Cy3-labelled Aβ42 monomer adsorption on a quartz substrate after L-DOPA coating by confocal fluorescence spectroscopy and molecular dynamic simulation showed the huge influence of Aβ42 adsorption on the aggregation process.
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Domaine public
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