  Two different metrics are used to assess Forster resonance energy transfer
(FRET) between fluorophores in the steady state: (1) acceptor-quenching of
donor fluorescence, E (a.k.a. transfer efficiency); and (ii) donor-excited
acceptor fluorescence, F-A-Dex. While E is still more widely used, F-A-Dex has
been gaining in popularity for practical reasons among experimentalists who
study biomembranes. Here, for the special case of membrane-bound fluorophores,
we present a substantial body of experimental evidence that justifies the use
of simple Stern-Volmer expressions when modeling either FRET metric under
dilute-probe conditions. We have also discovered a dilute-regime correspondence
between our Stern-Volmer expression for E and Wolber and Hudson's series
approximation for steady-state Forster quenching in 2D. This novel
correspondence allows us to interpret each of our 2D quenching constants in
terms of both (i) an effective Forster distance, and (ii) two maximum
acceptor-concentration limits, each of which defines its own useful
experimental regime. Taken together, our results suggest a three-step strategy
toward designing more effective steady-state FRET experiments for the study of
biomembranes.
