Table 1.

Advantages and disadvantages of different luminescent detection methods for DNA-binding proteins

Methods	Advantages	Disadvantages
Fluorescence anistropy or early FRET methods	Only one fluorescent label on the DNA is required.	Fluorescence polarization has a small dynamic range and depends on the sizes of the DNA and protein molecules. FRET signal depends on the number and position of endogenous fluorescent residues. Lack of generalizability.
MB strategy	An increased dynamic range not limited to the size of molecules or the number and position of endogenous fluorescent residues. Can be adapted for sensing different DNA-binding proteins by modification of the oligonucleotide binding site.	Limited to the detection of proteins with intrinsic DNA-binding activities. Labeling can be quite expensive and interfere with the protein–DNA interaction.
Aptamer beacon strategy	High affinity aptamers can be raised, in principle, for any protein target. Can be adapted for sensing different proteins by modification of the aptamer sequence.	Labeling can be quite expensive and interfere with the protein–DNA interaction. Some designs rely on specific aptamer structural features, reducing generalizability.
Label-free strategy	Economical, suitable for high-throughput or low-cost applications.	Cannot control the position of the luminescent dye on the oligonucleotide. Lack of multiplex detection capability.
Exonuclease protection strategy	No need to consider the equilibria between multiple oligonucleotide conformations and the bound and free states of the beacon or aptamer.	Can be more susceptible to false positives.