Research
New Surface Design Makes QDs More Reliable for Single-Molecule Imaging
Abstract Quantum dots (QDs) are fluorescent nanoparticles widely used for single-molecule imaging because of their exceptional brightness and photostability. However, the impact of QD surface chemistry on biomolecular interactions has not been systematically investigated. Here, we report that commercial QDs unexpectedly destabilize protein-DNA complexes by inducing protein dissociation from DNA. Using the human nucleotide excision repair protein, xeroderma pigmentosum complementation group A (XPA) as a model system, we demonstrate that antibody-conjugated QDs promote dissociation of XPA from DNA substrates, independently of sizes and surface modification of QDs, antibody types, epitope tags, buffer conditions, or DNA structures. We find that polyethylene glycol (PEG), a common polymer coating on QD surfaces, is the primary factor responsible for this effect. To tackle this problem, we engineered QDs with precisely controlled surface polymer compositions. By systematically changing the ratio of anchoring, hydrophilic, and PEG-based functional groups, we find that reducing PEG density below a critical threshold effectively suppresses protein dissociation while maintaining excellent colloidal stability and brightness. Furthermore, antibodies conjugated via click chemistry between azide groups and DBCO enabled specific labeling of XPA without perturbing the DNA binding activity. Using these optimized QDs, we conducted single-molecule DNA curtain assays to visualize XPA-DNA interactions. QD-labeled XPA exhibits one-dimensional diffusion with frequent pausing on undamaged DNA. DNA curtain assays revealed that XPA preferentially binds DNA bubbles and searches for bubble structures through both one-dimensional diffusion and three-dimensional collision. Quantitative analysis showed that three-dimensional collision is the dominant pathway for bubble recognition. Taken together, our results uncover a previously unrecognized limitation of PEG-coated QDs in single-molecule studies and provide an improved surface-engineering strategy to preserve native protein-DNA interactions. Newly engineered QDs establish robust platforms for accurate single-molecule visualization of biomolecular processes. Quantum dots (QDs) are widely used as fluorescent probes for single-molecule imaging because they are brighter and more photostable than conventional fluorescent dyes. Researchers at UNIST have now discovered that the surface chemistry of many commercially available QDs can unintentionally disrupt the protein–DNA interactions they are intended to observe. The team also established a surface-engineering strategy that minimizes this interference without compromising their imaging performance. Led by Professor Jaeil Lee of the Department of Biological Sciences and Professor Jongnam Park of the Department of Energy and Chemical Engineering, the researchers identified polyethylene glycol (PEG), a polymer widely used to coat commercial QDs, as the source of the interference. When PEG occupied too great a proportion of the particle surface, protein–DNA interactions became destabilized, revealing a previously unrecognized limitation of a widely used imaging probe. The finding arose while the team was investigating xeroderma pigmentosum complementation group A (XPA), a DNA repair protein involved in nucleotide excision repair. As the concentration of commercially available QDs increased, XPA unexpectedly dissociated from DNA. The same behavior persisted regardless of quantum dot size, antibody type, surface modification, buffer conditions, or DNA structure, indicating that the disruption originated from the probe itself rather than the biological system. To address this limitation, the researchers systematically redesigned the quantum dot surface by adjusting the proportions of anchoring, hydrophilic, and PEG-derived functional groups. They found that reducing the PEG-derived surface composition below approximately 7% preserved native protein–DNA interactions while maintaining the brightness and colloidal stability required for single-molecule imaging. Using the redesigned QDs, the researchers visualized how XPA searches for sites of DNA damage without perturbing its natural behavior. Single-molecule DNA curtain experiments showed that XPA locates DNA bubble structures through both one-dimensional diffusion along DNA and three-dimensional collisions from solution. Quantitative analysis revealed that three-dimensional collisions are the dominant pathway for recognizing damaged DNA under physiologically relevant protein concentrations. “Commercial quantum dots have long been regarded as passive fluorescent probes,” said Professor Lee. "Our study shows that their surface chemistry can directly influence the molecular interactions being observed. By identifying the source of this interference, we established a practical strategy for preserving native protein–DNA interactions during single-molecule imaging." Professor Park added, "Careful control of surface polymer composition allowed us to reduce molecular interference without compromising brightness or colloidal stability. We expect these design principles to support the development of more reliable nanoparticle-based probes for biological imaging." The study also included contributions from Youngseo Kim of the Department of Biological Sciences and Hyerim Kim of the Department of Energy and Chemical Engineering at UNIST. The findings were published online in Nano Convergence on June 3, 2026. The research was supported by the National Research Foundation of Korea (NRF), the Institute for Basic Science (IBS), and the Ministry of Trade, Industry and Energy (MOTIE). Journal Reference Youngseo Kim, Hyerim Kim, Munryul Choi, et al ., "Engineering quantum dot surfaces to preserve protein-DNA interactions for single-molecule visualization," Nano Converg ., (2026).