Frster Resonance Energy Transfer FRET Microscopy
Förster Resonance Energy Transfer (FRET) microscopy is a powerful imaging technique used to study molecular interactions and protein dynamics in living cells.
Principle of FRET
FRET is a non-radiative energy transfer process that occurs between two closely spaced fluorophores. When the excitation wavelength of the donor fluorophore overlaps with the absorption spectrum of the acceptor fluorophore, the donor fluorophore transfers its energy to the acceptor, causing the acceptor fluorophore to emit fluorescence.
Applications of FRET Microscopy
- Study of protein-protein interactions
- Investigation of protein conformational changes
- Monitoring of cellular signaling pathways
- Detection of protein localization and trafficking
Methods of FRET Microscopy
- Steady-state FRET: Measures the ratio of donor to acceptor fluorescence emission intensities under continuous illumination.
- Time-resolved FRET: Measures the decay time of donor fluorescence after pulsed excitation to determine the efficiency of FRET.
- Fluorescence lifetime imaging microscopy (FLIM): Creates images based on the fluorescence lifetime of the donor fluorophore, providing spatial information about FRET.
Advantages of FRET Microscopy
- Allows for real-time imaging of molecular interactions in live cells.
- Provides quantitative information about the distance and orientation of fluorophores.
- Can be combined with other imaging techniques for comprehensive cellular analysis.