Analyzing Amyloid Structures in a Tissue Section Using Fluorescence Lifetime Imaging Microscopy

Abstract

Source: Nyström, S., et al., Imaging Amyloid Tissues Stained with Luminescent Conjugated Oligothiophenes by Hyperspectral Confocal Microscopy and Fluorescence Lifetime Imaging. J. Vis. Exp. (2017).

This video demonstrates analyzing amyloid structures with fluorescence lifetime imaging microscopy or FLIM, using hFTAA dye to reveal compact cores as blue/green and unstable peripheries as red/yellow in color-coded images, reflecting amyloid organization.

Protocol

1. Preparation of Tissue Samples

NOTE: Many tissue types can be imaged using heptamer-formyl thiophene acetic acid (hFTAA) as an amyloid marker. hFTAA is sensitive to aggregate conformation. The staining should, hence preferably, be performed on undisrupted aggregates with no epitope exposure. Optimal spectral quality is achieved if the tissue fixation is kept to a minimum. Hence, fresh frozen material, gently fixed in ethanol at the time of staining, is preferred. However, it is possible to detect am....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
hFTAA/Amytracker545	Ebba Biotech
Dako fluorescene mounting medium	Agilent technologies	GM304
LeicaDM6000	Leica
Lumen 200	Prior
Spectraview system	ASI spectral imaging
Spectraview software	ASI spectral imaging
LSM780	Zeiss
Zen 2010b v6.0 software	Zeiss
FLIM system	Becker & Hickl
Ar/ML 458/488/514	Zeiss
Tunable Laser In Tune	Zeiss