Folding of nanostars was assessed using agarose gel electrophoresis (AGE) and dynamic light scattering (DLS). For these experiments the cholesterol-modified strand was replaced with a non-functionalized oligonucleotide of identical sequence to prevent aggregation. Mixtures of required oligonucleotides were prepared in Eppendorf tubes to give a final nanostar concentration of 10 µM. Prepared mixtures were cooled from 95 • C to 20 • C at -0.05 • C min -1 using a Techne TM TC-512 thermocycler to enable nanostructure hybridization. Annealed samples were stored at 4 • C and used for electrophoresis and light scattering studies within a week. Agarose gels were prepared at 1.5 wt% agarose in Tris-Borate-EDTA buffer (Sigma Aldrich), and precast with DNA stain (SYBR Safe, Ther-moFisher). Annealed nanostars were mixed with loading dye (Thermo Scientific) and a volume equivalent to 7.5 µg of DNA was loaded into each well. A potential of 75 V (3 V cm -1 ) was applied for 120 minutes. Gels were imaged using a GelDoc-It imaging system equipped with a UV lamp for illumination and CCD camera for image acquisition. DLS measurements were performed on a Malvern Zetasizer NanoZSP, equipped with a 633nm He-Ne laser with a maximum power of 10mW. The scattering angle was fixed at 173 • . An ultra low volume quartz cuvette (ZEN2112, Malvern) was filled with 50 µL of sample syringe-filtered through 0.22 µm polyvinylidene fluoride filters (Millex), and sealed with a polypropylene cap to prevent evaporation. Size distributions shown in Fig. are averaged over 3 runs, each of which consisted of 13-16 individual measurements. A Peltier heating block was used for temperature control.