Microfluidic-Generated Seeds for Gold Nanotriangle Synthesis in Three or Two Steps.

Nanoparticle synthesis has drawn great attention in the last decades. The study of crystal growth mechanisms and optimization of the existing methods lead to the increasing accessibility of nanomaterials, such as gold nanotriangles which have great potential in the fields of plasmonics and catalysis. To form such structures, a careful balance of reaction parameters has to be maintained. Herein, a novel synthesis of gold nanotriangles from seeds derived with a micromixer, which provides a highly efficient mixing and simple parameter control is reported. The impact of the implemented reactor on the primary seed characteristics is investigated. The following growth steps are studied to reveal the phenomena affecting the shape yield. The use of microfluidic seeds led to the formation of well-defined triangles with a narrower size distribution compared to the entirely conventional batch synthesis. A shortened two-step procedure for the formation of triangles directly from primary seeds, granting an express but robust synthesis is further described. Moreover, the need for a thorough study of seed crystallinity depending on the synthesis conditions, which - together with additional parameter optimization - will bring a new perspective to the use of micromixers which are promising for scaling up nanomaterial production is highlighted.

Flow imaging microscopy as a novel tool for high-throughput evaluation of elastin-like polymer coacervates.

Biological and bioinspired polymer microparticles have broad biomedical and industrial applications, including drug delivery, tissue engineering, surface modification, environmental remediation, imaging, and sensing. Full realization of the potential of biopolymer microparticles will require methods for rigorous characterization of particle sizes, morphologies, and dynamics, so that researchers may correlate particle characteristics with synthesis methods and desired functions. Toward this end, we evaluated biopolymer microparticles using flow imaging microscopy. This technology is widely used in the biopharmaceutical industry but is not yet well-known among the materials community. Our polymer, a genetically engineered elastin-like polypeptide (ELP), self-assembles into micron-scale coacervates. We performed flow imaging of ELP coacervates using two different instruments, one with a lower size limit of approximately 2 microns, the other with a lower size limit of approximately 300 nanometers. We validated flow imaging results by comparison with dynamic light scattering and atomic force microscopy analyses. We explored the effects of various solvent conditions on ELP coacervate size, morphology, and behavior, such as the dispersion of single particles versus aggregates. We found that flow imaging is a superior tool for rapid and thorough particle analysis of ELP coacervates in solution. We anticipate that researchers studying many types of microscale protein or polymer assemblies will be interested in flow imaging as a tool for quantitative, solution-based characterization.

Preparation and atomic force microscopy of quadruplex DNA.

The purpose of this chapter is to provide detailed instructions for the preparation and atomic force microscopy (AFM) imaging of linear chains of quadruplex DNA (a.k.a. "G-wire DNA"). Successful self-assembly of long chain quadruplex DNA requires pure concentrated guanine-rich oligonucleotide sequence (GROs) and monovalent cations in a growth buffer. AFM imaging of individual G-wire DNA strands requires many carefully monitored steps, including substrate preparation, G-wire concentration, adsorption onto substrate, rinsing, drying, appropriate selection/use of imaging probes, and dry atmosphere imaging conditions. Detailed step-wise instructions are provided.

A stable G-quartet binds to a huntingtin protein fragment containing expanded polyglutamine tracks.

Huntington's disease (HD) is a progressive neurodegenerative disorder that is inherited in an autosomal dominant fashion. The disease is the result of an expanded CAG repeat in exon 1 of the HD gene, which encodes an elongated polyglutamine tract in the mutant form of the protein, huntingtin. Disease pathogenesis is linked to intracellular aggregates that form because of the tendency of the mutant protein to misfold. The role of huntingtin aggregates in disease pathology is unclear; it has been proposed that the aggregates themselves are toxic because of their ability to sequester intracellular proteins and disrupt normal cellular function. In addition, the mechanistic steps that lead to aggregate formation appear to be central to HD pathology. We have previously reported that guanosine-rich oligonucleotides with the ability to fold into a G-quartet are effective inhibitors of the aggregation process of a huntingtin protein fragment with an elongated polyglutamine tract, Htn 1-171(Q58). The most active molecule is composed of 20 guanosine residues, which adopt a G-wire conformation. Here we establish that G20 inhibits protein aggregation as judged by native gel electrophoresis, an agarose gel electrophoresis for resolving aggregates (AGERA) assay, and an immunoblotting assay. We also visualize the G20-Htn1-171(Q58) protein complex by using a streptavidin-biotin pull-down assay as well as atomic force microscopy (AFM). The G20 molecule also interacts with Htn1-171(Q23), a fusion protein that contains 23 glutamine residues instead of 58 (Q58), but in a more degenerate and nonspecific fashion. Taken together, our data support the notion that G20 exhibits some selectivity in binding to specific protein species that assemble along the aggregation pathway.

Auto-orientation of G-wire DNA on mica.

Scanning probe microscopy was used to examine the orientation of Tet1.5 quadruplex DNA polymers, a.k.a. "G-wires", after adsorption onto freshly cleaved Phyllosilicate micas. The G-wires appear to have a preferential orientation at 60 degrees intervals after thorough rinsing and slow drying. The angles the G-wires made with the fast scan direction of the SPM probe were measured and the frequency-angle information was quantitatively characterized by an empirical correlation coefficient. Careful measurements indicate the Tet1.5 G-wires orient along the b lattice vector of mica, the next nearest neighbor potassium vacancy. A model is proposed to explain this auto-orientation affect due to alignment of the G-wires' phosphate backbone through magnesium tether cations. Pairs of adjacent, parallel phosphate groups of the G-wires (0.95 nm apart) appear to align with the next nearest neighbor potassium vacancy sites of mica (0.90 nm apart). This behavior is not observed in solution. The potential for using the auto-orientation phenomena in the development of high-density biomolecular nano-electronic devices is explored.