Figure 4.

Two common oligonucleotide-based strategies for isothermal amplication of fluorescence signal. (A) Hybridization chain reaction^[152] for enabling a FRET-based amplified fluorescence readout. The presence of a target sequence initiates a cascade hybridization reaction between two metastable hairpins (H1 and H2) with extended regions of complementarity. Incorporation of a FRET-pair, one in the loop region of H1 and the other in the toehold region of H2 brings the FRET-pair into close proximity upon HCR. (B) Hairpin DNA cascade amplification.^[72] Target binding to H1 exposes a region in H1 that can bind to H2. This releases the target and allows it to bind more H1 strands for amplified signal. The H1-H2 complex can bind to a fluorophore-labeled strand that is pre-hybridized to a quencher-labeled strand, displacing the latter and turning on fluorescence.