Time-Gated FRET Detection for Multiplexed Biosensing

Förster resonance energy transfer (FRET) has become an indispensable technique for biosensing in both microscopy and spectroscopy. FRET based on photoluminescence intensity and lifetime is both well known. However, their combination into time-gated (TG) intensity detection has been much less exploited, although many commercial diagnostic/screening assays and academic studies concerning TG FRET have demonstrated the successful application since more than two decades. In this chapter we explain the theoretical and practical background of TG FRET and discuss various applications. In particular, multiplexed detection of biomolecules and/or biomolecular interactions using photoluminescence spectroscopy and microscopy and the advantages and limitations of TG FRET compared to other technologies will be discussed.