PT  - JOURNAL ARTICLE
AU  - Chyi Wei Chung
AU  - Amberley D. Stephens
AU  - Edward Ward
AU  - Yuqing Feng
AU  - Molly Jo Davis
AU  - Clemens F. Kaminski
AU  - Gabriele S. Kaminski Schierle
TI  - Label-free characterisation of amyloids and alpha-Synuclein polymorphs by exploiting their intrinsic fluorescence property
AID  - 10.1101/2021.11.30.470691
DP  - 2021 Jan 01
TA  - bioRxiv
PG  - 2021.11.30.470691
4099  - http://biorxiv.org/content/early/2021/12/01/2021.11.30.470691.short
4100  - http://biorxiv.org/content/early/2021/12/01/2021.11.30.470691.full
AB  - Conventional in vitro aggregation assays often involve tagging with extrinsic fluorophores which can interfere with aggregation. We propose the use of intrinsic amyloid fluorescence lifetime represented by model-free phasor plots, as a label-free assay to characterise amyloid structure. Intrinsic amyloid fluorescence arises from structured packing of β-sheets in amyloids and is independent of aromatic-based fluorescence. We show that different amyloids (i.e., α-Synuclein (αS), β-Lactoglobulin and TasA) and different polymorphic populations of αS (induced by aggregation in salt-free and salt buffers mimicking the intra-/extracellular environments) can be differentiated by their unique fluorescence lifetimes. Moreover, we observe that disaggregation of pre-formed fibrils of αS and βLG leads to increased fluorescence lifetimes, distinct to those of their fibrillar counterpart. Our assay presents a medium-throughput method for rapid classification of amyloids and their polymorphs (the latter of which recent studies have shown lead to different disease pathology), and for testing small molecule inhibitory compounds.Competing Interest StatementThe authors have declared no competing interest.