Using live FRET imaging to reveal early protein-protein interactions during T cell activation

The emerging challenge for proteomics in general and lymphocyte biology in particular is to understand protein-protein interactions in the dynamic context of the living cell. Particularly interesting are the molecular dynamics of the T cell receptor-CD3 complex and other immunoreceptors in immune synapses. Fluorescence (or Forster) resonance energy transfer (FRET) is one of the few techniques that are capable of giving dynamic information about the nanometer-range proximity between molecules, as opposed to simply the subcellular co-localization that is provided by fluorescence microscopy. Spectral changes in fluorescence intensity and down modulation of donor lifetime are the basis for rapidly developing approaches to real-time FRET imaging. With two-photon excitation, FRET can now be extended to in vivo imaging.