Spectral FRET

Characterization of spectral FRET imaging microscopy for monitoring nuclear protein interactions

  • The spectral processed Forster resonance energy transfer ¨ (psFRET) imaging method provides an effective and fast method for measuring protein–protein interactions in living specimens. The commercially available linear unmixing algorithms efficiently remove the contribution of donor spectral bleedthrough to the FRET signal. However, the acceptor contribution to spectral bleedthrough in the FRET image cannot be similarly removed, since the acceptor spectrum is identical to the FRET spectrum. Here, we describe the development of a computer algorithm that measures and removes the contaminating ASBT signal in the sFRET image...