Fluorescence Lifetime Imaging of PolyQ Protein Aggregation in Caenorhabditis elegans Neurons

Begin with anesthetized, age-matched control and chaperone-deficient Caenorhabditis elegans worms. 

Both groups express fluorophore-tagged polyQ proteins in their neurons that are prone to misfolding. 

In control worms, chaperone proteins limit misfolded polyQ aggregation, whereas, in chaperone-deficient worms, misfolded polyQ readily forms aggregates. 

Place the control slide on a confocal microscope stage for Fluorescence Lifetime Imaging Microscopy (FLIM). 

Illuminate the worms using a pulsed laser. 

The laser excites the fluorophore, which emits a photon as it returns to the ground state. 

FLIM measures fluorescence lifetime—the duration the fluorophore remains in its excited state.

In chaperone-deficient worms, polyQ aggregation clusters fluorophores, promoting energy transfer between adjacent fluorophores instead of photon emission, thereby reducing fluorescence lifetime.   

The FLIM software processes these lifetimes to generate color-coded maps, enabling aggregation visualization.

A reduced fluorescence lifetime in chaperone-deficient worms, compared to controls, indicates increased polyQ aggregation.