Single-Molecule DNA Hybridization on Tetrahedral DNA Framework-Modified Surfaces

Tetrahedral DNA frameworks (TDFs) have been extensively
utilized
in biosensing systems. At the macroscopic level, it has been well
established that TDFs, when employed as probe scaffolds, significantly
enhance the interfacial DNA hybridization efficiency between target
single-stranded DNA (ssDNA) and probe ssDNA at the solid–liquid
interface. However, a molecular-level understanding of how TDF scaffolds
facilitate this interfacial hybridization remains elusive. In this
study, we employed single-molecule total internal reflection fluorescence
microscopy (SM-TIRFM) to monitor the hybridization processes between
freely diffusing target ssDNA and probe ssDNA with or without TDF
scaffolds. Statistical analysis of interfacial hybridization kinetics
at single probe sites revealed that TDF scaffolds significantly accelerate
the interfacial hybridization kinetics, reducing the target dissociation
time by 0.5-fold and increasing the association constant (Ka) by nearly 4-fold. This study offers new insights
for the development of DNA nanostructure-based biosensors.