Sep 17, 2009

The thioflavin T fluorescence assay for amyloid fibril detection can be biased by the presence of exogenous compounds

The FEBS Journal
Sean A HudsonJohn A Carver

Abstract

Thioflavin T (ThT) dye fluorescence is used regularly to quantify the formation and inhibition of amyloid fibrils in the presence of anti-amyloidogenic compounds such as polyphenols. However, in this study, it was shown, using three polyphenolics (curcumin, quercetin and resveratrol), that ThT fluorescence should be used with caution in the presence of such exogenous compounds. The strong absorptive and fluorescent properties of quercetin and curcumin were found to significantly bias the ThT fluorescence readings in both in situ real-time ThT assays and single time-point dilution ThT-type assays. The presence of curcumin at concentrations as low as 0.01 and 1 mum was sufficient to interfere with the ThT fluorescence associated with fibrillar amyloid-beta(1-42) (0.5 mum) and fibrillar reduced and carboxymethylated kappa-casein (50 mum), respectively. The ThT fluorescence associated with fibrillar amyloid-beta(1-42) was also biased using higher concentrations of resveratrol, a polyphenol that is not spectroscopically active at the wavelengths of ThT fluorescence, implying that there can be direct interactions between ThT and the exogenous compound and/or competitive binding with ThT for the fibrils. Thus, in all cases where ThT i...Continue Reading

  • References30
  • Citations160

Mentioned in this Paper

APP protein, human
Kappa-Caseins
UniProtKB
Fluorescence Spectroscopy
Fluorescent stain
Dietary Polyphenol
Electron Microscopy, Diagnostic
Amyloid Deposition
Thioflavin T
Electron Microscopy

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