Aggregation of misfolded proteins leading to the formation of the so-called amyloid fibrils is a feature of many neurodegenerative disorders, such as Parkinson’s, Alzheimer’s and Huntington’s diseases. The ability to resolve these fibrils in vitro or in situ is therefore a key requirement to understand the molecular mechanisms of these diseases and potentially progress in the search for therapeutic intervention. However, the characteristic length scale of these protein assembly is two orders of magnitude smaller than the spatial resolution of conventional optical microscopes. This suggests that optical super-resolution microscopy techniques, which have been awarded by the Nobel Prize in Chemistry 2014, have the potential to study their formation.

In this context, I will report on the implementation and application of a Stimulated Emission Depletion microscope (STED) [3] to monitor the self-assembly of misfolded proteins. Our home-built STED microscope is based on several advanced technological solutions such as a supercontinuum source and a spatial light modulator. The former is a simple and cost effective component to excite various dyes, while the latter provides an effective way to generate aberration-free depletion patterns and co-align perfectly the excitation and depletion beams without moving any mechanical parts.

I will then present the use of STED microscopy to study the formation of α-synuclein fibrils in vitro and a method to extract quantitative parameters on the fibril kinetics from super resolved images. Eventually, I will report on the combination of STED microscopy with Atomic Force Microscopy (AFM) [4]. Interestingly, STED microscopy enables the fast and specific localisation of labelled aggregates on large field of view while AFM provides a locally better resolved morphological map and potentially insight into the mechanical properties of the aggregates.


Wednesday, January 21, 2015, 12:00. Seminar Room

Hosted by Dr. Pablo Loza