Depending on the nature of the study and what sort of information one is trying to gather through the use of FRET, FLIM has some compelling advantages in certain situations, and can provide a quantitative evaluation of the donor, acceptor and FRET pair stoichiometry. It does require access to specialized equipment and software. Different approaches to FLIM data acquisition have different strengths and weaknesses. For dynamic studies requiring high time resolution, FLIM acquisition times can fall well short of ideal.

If a yes/no answer to whether FRET is occurring is all that is required, then the polarization anisotropy of the acceptor can be used to determine FRET between fluorescent proteins (Rizzo and Piston, 2005). This is a relatively simple and robust method for confirming the presence/absence of FRET.