Walawalkar, V. | Search bioRxiv

bioRxiv2023

Membrane and glycocalyx tethering of DNA nanostructures for enhanced uptake

DNA nanostructures (DNs) have been increasingly utilized in biosensing, drug delivery, diagnostics and therapeutics, because of their programmable assembly, control over size and shape, and ease of functionalization. However, the low cellular uptake of DNs has limited their effectiveness in these biomedical applications. Here we demonstrate the potential of membrane and glycocalyx binding as general strategies to enhance the cellular uptake of DNs. By targeting the plasma membrane and cell-surface glycocalyx, the uptake of all three distinct DNs is significantly enhanced as compared to uptake of bare DNs. We also demonstrate the viability of single-step membrane labeling by cholesterol-DNs as competitive with previous multistep approaches. Further, we show that the endocytic pathway of membrane-bound DNs is an interdependent process that involves scavenger receptors, clathrin-, and caveolinmediated endocytosis. Our findings may potentially expand the toolbox for effective cellular delivery of DNA nanostructured systems.

bioengineering↗

bioRxiv2023

Capacitance measurements for assessing DNA origami nanostructures

Nanostructures fabricated with DNA are emerging as a practical approach for applications ranging from advanced manufacturing to therapeutics. To support the strides made in improving accessibility and facilitating commercialization of DNA nanostructure applications, we identify the need for a rapid characterization approach that aids nanostructure production. In our work, we introduce a low-fidelity characterization approach that provides an interdependent assessment of DNA origami formation, concentration and morphology using capacitance sensing. Change in charge is one of the transduction methods to determine capacitive loading on a substrate. It is known that cations in the solution stabilize DNA origami nanostructures. So, we hypothesized that the presence of cations and nanostructures in a buffer solution can induce capacitance change that is distinctive of the nanostructure present. In this study we were able to detect a change in the capacitance when the nanostructure solution was deposited on our capacitance sensor, and we could distinguish between pre-annealed and annealed structures at concentrations less than 15 nM. The capacitance measurements were affected by the concentration of Mg2+ ions in the solution, the staple-to-scaffold stoichiometric ratio of the nanostructure and the nanostructure morphology. Maintaining a 12.5 mM Mg2+ concentration in the nanostructure buffer, we discover a linear relationship between the relative capacitance change and the nanostructure concentration from 5 nM to 20 nM, which we call the characteristic curve. We find distinct characteristic curves for our three nanostructures with distinct morphologies but similar molecular weight - a rectangular plate, a sphere and a rod. Given that we can distinguish nanostructure formation, concentration and morphology, we expect that capacitance measurement will emerge as an affordable and rapid approach for quality control for nanostructure production.

bioengineering↗