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AssigneeThe University Court of the University of Edinburgh
InventorsAlison Green, Graham Fairfoul
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A method of detecting the presence of alpha-synuclein aggregation in a biological sample is provided whereby a biological sample is mixed with a reaction sample comprising a population of beads, a fluorophore adapted to hind to protein aggregates and to increase fluorescence when bound to protein aggregates, and alpha-synuclein or a fragment or variant thereof to forum a reaction mixture, the reaction mixture is illuminated and at the same time incubated with intermittent agitation cycles, wherein a significant increase in the fluorescence of the reaction mixture during incubation is indicative of the presence of aggregates of alpha-synuclein in the biological sample. Method of diagnosing alpha-synucleinopathies such as Parkinson's disease or Dementia with Lewy Bodies.
A method of detecting the presence of alpha-synuclein aggregation in a biological sample, the method comprising the steps:
The method according to any one preceding claim, wherein the reaction sample is a buffered reaction sample.
The method according to any one preceding claim, wherein the protein aggregates comprise significant beta-sheet content.
The method according to any one preceding claim, wherein the fluorophore is a thioflavin or cyanine T-284.
The method according to any one preceding claim, wherein the reaction sample comprises from approximately 0.01 mg/mL alpha-synuclein to about 10 mg/mL alpha-synuclein to act as an aggregation substrate.
The method according to any one preceding claim, wherein the alpha-synuclein of the reaction sample is a fragment of full length alpha-synuclein.
The method according to any one preceding claim, wherein the beads of the population of beads of the reaction sample comprise zirconia, silica, glass, quartz, or a polymer such as polystyrene, polytetrafluoroethylene (PTFE), polymethylmethacrylate (PMMA), or combinations thereof.
The method according to any one preceding claim, wherein the population of beads have a mean diameter of the beads from approximately 1 mm to approximately 0.001 mm.
The method according to any one preceding claim, wherein the reaction sample comprises from approximately 1 mg to approximately 150 mg of beads per 100 μL of reaction mixture.
The method according to any one preceding claim, wherein the presence of aggregates of alpha-synuclein in the biological sample is indicative of a disease associated with the abnormal aggregation of alpha-synucein such as Dementia with Lewy Bodies or Parkinson's Disease, or other alpha-synucleinopathies.
The method according to any one preceding claim, wherein the reaction mixture is incubated for more than 40 hours, more than 60 hours, more than 80 hours, or more than 120 hours.
The method according to any one preceding claim, wherein the method is carried out at a temperature of from 25° C. to 45° C.
The method according to any one preceding claim, wherein the biological sample is treated prior step (i) to concentrate the alpha-synuclein in the biological sample.
A method of diagnosing alpha-synucleinopathies, the method comprising the steps:
A kit of parts comprising an aqueous buffered solution, the aqueous buffered solution comprising a population of beads, a fluorophore and alpha-synuclein or a fragment or variant thereof.
Dependencies are inferred from the claim text.