Table 2 Comparison between BiFC and FRET.
| Â | BiFC | FRET |
|---|---|---|
Required microscopic equipment | simple | extensive |
Data analysis and computation required | - | + |
Concentration dependence | high | high (channel FRET, FSPIM) or low (DFRAP, FLIM) |
Specific problems | false positives (possibly due to high expression levels and/or irreversibility) | donor bleed-through (channel FRET, FSPIM), photoconversion, protein mobility (DFRAP) |
Endogenous expression control (i.e. visualization of tagged partners with subcellular resolution) | - | + |
Monitoring interaction dynamics | - (fluorophore reconstitution irreversible) | + (interactions reversible) |
Subcellular resolution of interaction sites | high | high (FLIM) or low (channel FRET, FSPIM or DFRAP) |
Suitable for tri-molecular interactions | - | +/- |
Suitable for monitoring multiple distinct interaction pairs inside the same cell ("multicolor") | + | - |
Suitable for medium to high throughput | + | - |