hFTAA Staining of Amyloid Deposits: A Technique to Visualize Amyloid Fibrils Using Oligothiophene-Based Dye for Fluorescence Imaging in Tissue Sections
DEŞİFRE METNİ
The deposition of misfolded protein aggregates – the amyloids in the extracellular tissue space are hallmarks of various proteopathic diseases. These amyloids are composed of beta sheets which align perpendicular to the fibrillar axis, forming cross beta-sheet repeats.
To detect the amyloid deposits with heptamer-formyl thiophene acetic acid, or hFTAA, stain, take a formalin-fixed, paraffin-embedded tissue section containing amyloid deposits.
Treat the sections with xylene to deparaffinize the tissue for improving the accessibility to stain in the subsequent steps. Add a few drops of the hFTAA staining solution over the tissue section and incubate to allow the dye to penetrate the tissue.
The hFTAA contains seven conjugated sulfur-containing thiophene units and terminal carboxyl groups attached to the thiophene backbone. These features provide flexibility to the unbound dye. During incubation, the end carboxyl groups of the dye bind with the positively charged amino acid residues lining the hydrophobic groove of the beta-sheet via electrostatic interactions.
Consequently, a conformational change occurs that reduces the structural flexibility of the bound hFTAA dye. This enhances the bound dye’s fluorescence quantum yield – the relative ratio of absorbed versus emitted photons. This process helps to improve the visualization of the amyloids.
Observe the slide under a fluorescence microscope. The presence of yellow-red fluorescence spots indicates the presence of amyloids in tissue sections.
