Ultrasensitive Measurement of Ca(2+) Influx into Lipid Vesicles Induced by Protein Aggregates

Patrick Flagmeier, Suman De, David C Wirthensohn, Steven F Lee, Cécile Vincke, Serge Muyldermans, Tuomas P J Knowles, Sonia Gandhi, Christopher M Dobson, David Klenerman

Onderzoeksoutput: Articlepeer review

65 Citaten (Scopus)

Samenvatting

To quantify and characterize the potentially toxic protein aggregates associated with neurodegenerative diseases, a high-throughput assay based on measuring the extent of aggregate-induced Ca(2+) entry into individual lipid vesicles has been developed. This approach was implemented by tethering vesicles containing a Ca(2+) sensitive fluorescent dye to a passivated surface and measuring changes in the fluorescence as a result of membrane disruption using total internal reflection microscopy. Picomolar concentrations of Aβ42 oligomers could be observed to induce Ca(2+) influx, which could be inhibited by the addition of a naturally occurring chaperone and a nanobody designed to bind to the Aβ peptide. We show that the assay can be used to study aggregates from other proteins, such as α-synuclein, and to probe the effects of complex biofluids, such as cerebrospinal fluid, and thus has wide applicability.

Originele taal-2English
Pagina's (van-tot)7750-7754
Aantal pagina's5
TijdschriftAngewandte Chemie International Edition
Volume56
Nummer van het tijdschrift27
DOI's
StatusPublished - 26 jun 2017

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