The transfer of electronic excitation energy from small terbium complexes to chromophores in solution is greatly enhanced by the fact that during its millisecond luminescence lifetime a typical excited terbium complex diffuses extensively (covering a net distance of about 2 × 103 nm). Thus, under ordinary circumstances, an excited terbium complex actually encounters a large number of possible energy acceptors before it decays. (At 10-6 M, the average separation between neighboring acceptors is 65 nm.) This has the effect of averaging the interactions between energy donor and acceptor(s) over all allowed directions and orientations in space, so that the final result reflects the equilibrium properties of the system under study. Energy transfer in this rapid-diffusion limit is sensitive to the electric charge of the donor and acceptor. A set of terbium probes with the same size and approximately spherical shape but different charges (1+, 0, 1-) permits quantitative study of electrostatic effects. These have been calibrated by studies of energy transfer to small, spherical transition metal complexes. They have subsequently been employed in studies of chromophores in proteins, and DNA-binding drugs. Measurements of the effect of DNA on energy transfer between small, freely diffusing ions indicate how cations cluster in regions near DNA, and how anions are repelled from the same regions.
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