Characterization of DNA Origami Nanostructures for Size and Concentration by Dynamic Light Scattering and Nanoparticle Tracking Analysis

Nucleic acids self-assembly has rapidly advanced to produce multi-dimensional nanostructures with precise sizes and shapes. DNA nanostructures hold great potential for a wide range of applications, including biocatalysis, smart materials, molecular diagnosis, and therapeutics. Here, we present a study of using dynamic light scattering (DLS) and nanoparticles tracking analysis (NTA) to analyze DNA origami nanostructures for their size distribution and particles concentrations. Compared to DLS, NTA demonstrated higher resolution of size measurement with a smaller FWHM and was well suited for characterizing multimerization of DNA nanostructures. We future used intercalation dye to enhance the fluorescence signals of DNA origami to increase the detection sensitivity. By optimizing intercalation dyes and the dye-to-DNA origami ratio, fluorescent NTA was able to accurately quantify the concentration of dye-intercalated DNA nanostructures, closely matching with values obtained by UV absorbance at 260 nm. This optimized fluorescent NTA method offers an alternative approach for determining the concentration of DNA nanostructures based on their size distribution, in addition to commonly used UV absorbance quantification. This detailed information of size and concentration is not only crucial for production and quality control but could also provide mechanistic insights in various applications of DNA nanomaterials.