In recent years super-hydrophobic micro-patterned substrates (SHS) have been successfully used for the suspension of a few biological molecules, allowing the further characterization in a background-free environment by label-free techniques such as Raman spectroscopy, SEM and TEM in one device. This result is due to the combined action of laminar flow and shear stress exerted on the molecules contained in a drop that is spotted on top of the SHS and slowly evaporates. This new method is here proposed for the label-free formation and background-free characterization of amyloid fibers.
Amyloids are insoluble aggregates formed by proteins that convert from a misfolded form into highly-organized β-sheet structures that could accumulate in different organs and compromise their normal physiological functions. Known amyloid-related diseases, named amyloidosis, are for instance Alzheimer, Parkinson, and type 2 diabetes. In classical crystallography, the study of the amyloid aggregates structure is often hampered by the laborious and time consuming sample preparation techniques. Therefore the need of a quick reproducible technique, has emerged.
The amyloid fibers investigated in this work are derived from a lysozyme protein and a Tau-derived short peptide, both known to be related to two forms of amyloidosis. With this technique we demonstrate that threads of protein fibers are deposited on the substrate helped by the patterning of the SHS and its properties, and by characterizing them with Raman spectroscopy technique we revealed that they are anisotropic structures of amyloid nature. This type of sample preparation technique arises from the effect of the evaporation on the SHS, and opens up new possibilities for the formation of oriented fibers of amyloids and more in general, of proteins, ready for a substrate-free characterization, while classic crystallographic methods could have a limitation.
|Date of Award||Nov 22 2018|
|Original language||English (US)|
- Physical Sciences and Engineering
|Supervisor||Enzo Di Fabrizio (Supervisor)|
- Amyloid Fibers
- Single Device