Molecular beacons involving highly luminescent colloidal quantum dots

Abstract

Molecular beacons are fluorescently labeled biological systems, such as sequence-specific oligonucleotide hairpin structures with a stem and a loop, Eclipse probes, and nonnucleotide protein-based structures that react to ligand binding by generating a strongly enhanced fluorescent signal. For example, in oligonucleotide hairpin structures with a stem and a loop, a fluorescent probe (emitter) is chemically attached to one end of the loop structure, and a fluorescent quencher is attached to the other end.1 In the off-state, the fluorescent emitter is in close contact with the quencher, and luminescence is absent due to the Förster resonance energy transfer (FRET) or another type of energy transfer (ET) from the emitter to the quencher. During the detection procedure, the molecular beacon binds to its target, the complementary oligonucleotide sequence, opening the hairpin structure and spatially separating the emitter and the quencher, thus enabling luminescence. The detection scheme may be different if it is based on single-color fluorescence quenching or fluorescence release upon targeted binding to the analyte or spectrally encoded fluorescence signal, when both the FRET donor and the FRET acceptor are fluorescence probes capable of emitting. However, the main principle of the beacon operation is always the same and is based on detection of the changes in FRET efficiency between the emitter and the quencher via controllable spatial separation between the FRET donor and acceptor.