Alzheimer’s disease is the most common cause of dementia and was responsible for over 2% of all deaths in Sweden 2012. One of the pathological hallmarks is amyloid plaques built by fibrillated Amyloid β. Luminescent conjugated oligothiophenes are known to stain and give characteristic fluorescence spectra when staining amyloid fibrils. Little is however known about the interactions between LCOs and fibrils. Studies have been performed on molecules more traditionally known to stain amyloid fibrils. Studies have also been performed on fibrils using limited proteolysis. So far no studies have been performed using LCOs combined with limited proteolysis in order to study the interaction pattern between LCOs and fibrils. Amyloid β is expressed and purified using a simple few step purification protocol. The amyloid β peptide was then fibrillated in several generations in order to select for a homogenous fibril structure. This purification protocol also has the ability to purify different oligomers of Amyloid β that are interesting from a toxicity point of view. In this thesis optical characteristics and limited proteolysis with mass spectrometry are being used to studies the interactions between LCOs and fibrillated amyloid β. The proteolytic pattern was suggestive of an accessible N-terminal and a hidden C-terminal of Amyloid β M1-42 in the fibril. It was also shown that the proteolysis cleavage pattern of Chymotrypsin is not disrupted when the LCO pKTAA was used to stain fibrils. The emission spectra from the two LCOs pATAA and pKTAA changes differently when subjected to continuous excitation indicative of conformational changes or chemical modification.

Transthyretin (TTR) amyloidosis is a disease that appears in three variants. One variant affects the elderly population with heart failure, the other two variants are hereditary and caused by an amino acid substitution in the gene, resulting in polyneuropathy and/or heart issues depending on the amino acid substitution. However, in all three variants, other organs may also be affected with amyloid deposition in the disease course. Amyloid fibrils of TTR (ATTR) contains a mixture of full-length protein and fragments (50-127). Luminescent conjugated oligothiophenes (LCO’s) are novel amyloid binding probes used to stain amyloid fibrils and these amyloid probes have the feature of characterizing the amyloid structure in terms of fluorescence spectra. Apart from LCO’s, a few other amyloid binding probes are used to stain recombinant amyloid transthyretin and native transthyretin for binding studies. The majority of generated TTR aggregates in vitro did not have the characteristic fluorescence spectra when bound to LCO’s and was observed as a clumped gel-like aggregate. The generation of recombinant TTR fibrils in vitro using the mutant TTR-T49M to obtain an aggregation prone fragment (50-127) after being treated with cyanogen bromide had a low yield of in vivo amyloid-like fibrils, but with characteristic LCO spectra. Carpal tunnel ATTR often precedes ATTR deposition in heart tissue. Amyloid transthyretin in carpal tunnel tissues was stained with LCO’s and used as a reference in the comparison against the in vitro generated recombinant amyloid transthyretin fibrils. This project also includes quantification of amyloid transthyretin in a few selected parts of the carpal tunnel tissue using ImageJ. In the long run this method could help in diagnosing TTR amyloidosis.