DNA Linkers and Diluents for Ultrastable Gold Nanoparticle Bioconjugates in Multiplexed Assay Development

A novel bioconjugation strategy leading to ultrastable gold nanoparticles (AuNPs), utilizing DNA linkers and diluents in place of traditional self-assembled monolayers, is reported. The protective capacity of DNA confers straightforward biomolecular attachment and multistep derivatization capabilities to these nanoparticles and, more significantly, substantially enhances their stability in demanding and complex sensing environments. The DNA/AuNPs were assembled through pH-assisted thiol-gold bonding of single stranded DNA and salt aging, with preconjugated biotin moieties facing outward from the gold surface. These nanoparticles remain a stable colloidal suspension under a wide range of buffers and ionic strengths and can endure multiple rounds of lyophilization while retaining high biological activity. Furthermore, the high stability of the DNA/AuNPs allows for multiple reactions and conjugations to be performed within the colloidal suspensions (i.e., Protein A and antibody binding) for tailored and specific recognition to take place. We have demonstrated the applications of the DNA/AuNPs for colorimetric assays and ELISA feasibility; additionally, SPR imaging analysis of a supported membrane microarray shows excellent results with DNA/AuNPs as the enhancing agent. Together, the properties imparted by this interface render the material suitable for clinical and point-of-care applications where stability, throughput, and extended shelf lives are needed.