Colorimetric Biosensor for Nucleic Acid Detection Using Hybridization Chain Reaction and Gold Nanoparticles: Effect...

In this work, a pair of DNA hairpins was specifically designed for detecting and amplifying a target DNA via hybridization chain reaction (HCR). The biosensor exploits the interactions between gold nanoparticles (AuNPs) and DNA hairpins as well as HCR products, resulting in salt-induced aggregation (SIA) of the AuNPs, observable through visible color changes and UV-vis spectral shifts. This study builds on previous findings regarding the optimal length and sequence of hairpin toeholds and stems, which provide AuNPs with greater protection against SIA. The HCR-AuNP biosensor was developed by applying design guidelines for HCR hairpins and incorporating features that enhance their adsorption onto AuNPs. Intriguingly, hairpins with a 12-14 bp stem and a mixed-base toehold (6-8 nt) containing only a single adenine base provided adequate AuNP stabilization and enabled efficient HCR without leakage. The limit of detection (LOD) for the final HCR-AuNP biosensor was 10.7 nM, with the AuNP color changing from red to blue in the presence of the target DNA. This biosensor is advantageous due to its ease of operation, isothermal nature, and rapid results, all achieved without the need for enzymes or expensive equipment.