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Silver nanoclusters (AgNCs) are the next-generation nanomaterials representing supra-atomic structures where silver atoms are organized in a particular geometry. DNA can effectively template and stabilize these novel fluorescent AgNCs. Only a few atoms in size – the properties of nanoclusters can be tuned using only single nucleobase replacement of C-rich templating DNA sequences. A high degree of control over the structure of AgNC could greatly contribute to the ability to fine-tune the properties of silver nanoclusters. In this study, we explore the properties of AgNCs formed on a short DNA sequence with a C 12 hairpin loop structure (AgNC@hpC 12 ). We identify three types of cytosines based on their involvement in the stabilization of AgNCs. Computational and experimental results suggest an elongated cluster shape with 10 silver atoms. We found that the properties of the AgNCs depend on the overall structure and relative position of the silver atoms. The emission pattern of the AgNCs depends strongly on the charge distribution, while all silver atoms and some DNA bases are involved in optical transitions based on molecular orbital (MO) visualization. We also characterize the antibacterial properties of silver nanoclusters and propose a possible mechanism of action based on the interactions of AgNCs with molecular oxygen. 

Particle tracking (PT) microrheology is a passive microrheological approach that characterizes material properties of soft matter. Multicomponent materials with the ability to create extensive crosslinking, such as supra-assemblies, may exhibit a complex interplay of viscous and elastic properties with a substantial contribution of liquid phase still diffusing through the system. Microrheology analyzes the motion of microscopic beads immersed in a sample, making it possible to evaluate the rheological properties of biological supra-assemblies. This method requires only a small volume of the sample and a relatively simple, inexpensive experimental setup. The objective of this chapter is to describe the experimental procedures for the observation of particle motion, calibration of an optical setup for particle tracking, preparation of imaging chambers, and the use of image analysis software for particle tracking in viscoelastic nucleic acid-based supra-assemblies. 

The protocol described in this chapter allows for acquiring topography images of RNA-based nanoring structures and assessing their dynamic properties using atomic force microscopy (AFM) imaging. AFM is an indispensable tool for characterization of nucleic acid-based nanostructures with the exceptional capability of observing complexes in the range of a few nanometers. This method can visualize structural characteristics and evaluate differences between individual structurally different RNA nanorings. Due to the highly resolved AFM topography images, we introduce an approach that allows to distinguish the differences in the dynamic behavior of RNA nanoparticles not amenable to other experimental techniques. This protocol describes in detail the preparation procedures of RNA nanostructures, AFM imaging, and data analysis.