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Schematic representation of intermolecular terbium-to-GFP LRET. TMP-Lumi4 binds specifically and tightly to eDHFR (KD = ∼ 2 nM). Interaction between eDHFR and GFP fusion proteins and excitation of TMP-Lumi4 (the donor) results in LRET-sensitized emission of GFP (the acceptor). Time-resolved microscopy detects long-lifetime, sensitized GFP emission while eliminating cellular autofluorescence and directly excited GFP fluorescence background signals.
