Automated Ex Situ Assays of Amyloid Formation on a Microfluidic Platform
Yates, Emma V
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Saar, K., Yates, E. V., Müller, T., Saunier, S., Dobson, C., & Knowles, T. Automated Ex Situ Assays of Amyloid Formation on a Microfluidic Platform. Biophysical Journal, 110 555-560. https://doi.org/10.1016/j.bpj.2015.11.3523
Increasingly prevalent neurodegenerative diseases are associated with the formation of nanoscale amyloid aggregates from normally soluble peptides and proteins. A widely used strategy for following the aggregation process and defining its kinetics involves the use of extrinsic dyes that undergo a spectral shift when bound to β-sheet rich aggregates. An attractive route to carry out such studies is to perform ex situ assays, where the dye molecules are not present in the reaction mixture but rather are only introduced into aliquots taken from the reaction at regular time intervals in order to avoid the possibility that the dye molecules interfere with the aggregation process. However, such ex situ measurements are time-consuming to perform, require large sample volumes and do not provide for real-time observation of aggregation phenomena. To overcome these limitations, here we have designed and fabricated microfluidic devices that offer continuous and automated real-time ex situ tracking of the protein aggregation process. This device allows us to improve the time resolution of ex situ aggregation assays relative to conventional assays by more than one order of magnitude. The availability of an automated system for tracking the progress of protein aggregation reactions without the presence of marker molecules in the reaction mixtures opens up the possibility of routine non-invasive study of protein aggregation phenomena.
Financial support from the Frances and Augustus Newman Foundation, the BBSRC, the EPSRC, the ERC and the Swiss National Science Foundation is gratefully acknowledged.
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External DOI: https://doi.org/10.1016/j.bpj.2015.11.3523
This record's URL: https://www.repository.cam.ac.uk/handle/1810/253014
Attribution 2.0 UK: England & Wales
Licence URL: http://creativecommons.org/licenses/by/2.0/uk/
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