Absolute Quantification of Amyloid Propagons by Digital Microfluidics
2017
The self-replicating properties of proteins into amyloid fibrils is a common phenomenon and underlies a variety of neurodegenerative diseases. Because propagation-active fibrils are chemically indistinguishable from innocuous aggregates and monomeric precursors, their detection requires measurements of their replicative capacity. Here we present a digital amyloid quantitative assay (d-AQuA) with insulin as model protein for the absolute quantification of single replicative units, propagons. D-AQuA is a microfluidics-based technology that performs miniaturized simultaneous propagon-induced amplification chain reactions within hundreds to thousands of picoliter-sized droplets. At limiting dilutions, the d-AQuA reactions follow a stochastic regime indicative of the detection of single propagons. D-AQuA thus enables absolute quantification of single propagons present in a given sample at very low concentrations. The number of propagons quantified by d-AQuA was similar to that of fibrillar insulin aggregates d...
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