Controlling Non-Equilibrium Structure Formation on the Nanoscale.

Controlling the structure formation of gold nanoparticle aggregates is a promising approach towards novel applications in many fields, ranging from (bio)sensing to (bio)imaging to medical diagnostics and therapeutics. To steer structure formation, the DNA-DNA interactions of DNA strands that are coated on the surface of the particles have become a valuable tool to achieve precise control over the interparticle potentials. In equilibrium approaches, this technique is commonly used to study particle crystallization and ligand binding. However, regulating the structural growth processes from the nano- to the micro- and mesoscale remains elusive. Here, we show that the non-equilibrium structure formation of gold nanoparticles can be stirred in a binary heterocoagulation process to generate nanoparticle clusters of different sizes. The gold nanoparticles are coated with sticky single stranded DNA and mixed at different stoichiometries and sizes. This not only allows for structural control but also yields access to the optical properties of the nanoparticle suspensions. As a result, we were able to reliably control the kinetic structure formation process to produce cluster sizes between tens of nanometers up to micrometers. Consequently, the intricate optical properties of the gold nanoparticles could be utilized to control the maximum of the nanoparticle suspension extinction spectra between 525 nm and 600 nm.

Kinetically guided colloidal structure formation.

The self-organization of colloidal particles is a promising approach to create novel structures and materials, with applications spanning from smart materials to optoelectronics to quantum computation. However, designing and producing mesoscale-sized structures remains a major challenge because at length scales of 10-100 µm equilibration times already become prohibitively long. Here, we extend the principle of rapid diffusion-limited cluster aggregation (DLCA) to a multicomponent system of spherical colloidal particles to enable the rational design and production of finite-sized anisotropic structures on the mesoscale. In stark contrast to equilibrium self-assembly techniques, kinetic traps are not avoided but exploited to control and guide mesoscopic structure formation. To this end the affinities, size, and stoichiometry of up to five different types of DNA-coated microspheres are adjusted to kinetically control a higher-order hierarchical aggregation process in time. We show that the aggregation process can be fully rationalized by considering an extended analytical DLCA model, allowing us to produce mesoscopic structures of up to 26 µm in diameter. This scale-free approach can easily be extended to any multicomponent system that allows for multiple orthogonal interactions, thus yielding a high potential of facilitating novel materials with tailored plasmonic excitation bands, scattering, biochemical, or mechanical behavior.

Mechanics of soft epithelial keratin networks depend on modular filament assembly kinetics.

UNLABELLED: Structural adaptability is a pivotal requirement of cytoskeletal structures, enabling their reorganization to meet the cellular needs. Shear stress, for instance, results in large morphological network changes of the human soft epithelial keratin pair K8:K18, and is accompanied by an increase in keratin phosphorylation levels. Yet the mechanisms responsible for the disruption of the network structure in vivo remain poorly understood. To understand the effect of the stress-related site-specific phosphorylation of the K8:K18 pair, we created phosphomimicry mutants - K8(S431E), K8(S73E), K18(S52E) - in vitro, and investigated the various steps of keratin assembly from monomer to network structure using fluorescence and electron microscopy, and using rheology characterized their network mechanical properties. We find that the addition of a charged group produces networks with depleted intra-connectivity, which translates to a mechanically weaker and more deformable network. This large variation in network structure is achieved by the formation of shorter mutant filaments, which exhibit differing assembly kinetics and a manifestly reduced capacity to form the extended structures characteristic of the wild-type system. The similarity in outcome for all the phosphomimicry mutants explored points to a more general mechanism of structural modulation of intermediate filaments via phosphorylation. Understanding the role of kinetic effects in the construction of these cytoskeletal biopolymer networks is critical to elucidating their structure-function properties, providing new insight for the design of keratin-inspired biomaterials. STATEMENT OF SIGNIFICANCE: Structural remodeling of cytoskeletal networks accompanies many cellular processes. Interestingly, levels of phosphorylation of the human soft epithelial keratin pair K8:K18 increase during their stress-related structural remodeling. Our multi-scale study sheds light on the poorly understood mechanism with which site-specific phosphorylation induces disruption of the keratin network structure in vivo. We show how phosphorylation reduces keratin filament length, an effect that propagates through to the mesoscopic structure, resulting in the formation of connectivity-depleted and mechanically weaker networks. We determine that the intrinsically-set filament-to-filament attractions that drive bundle assembly give rise to the structural variability by enabling the formation of kinetically-arrested structures. Overall, our results shed light on how self-assembled intermediate filament structures can be tailored to exhibit different structural functionalities.

Assessing the ecological long-term impact of wastewater irrigation on soil and water based on bioassays and chemical analyses.

The reuse of treated wastewater for irrigation and groundwater recharge can counteract water scarcity and reduce pollution of surface waters, but assessing its environmental risk should likewise consider effects associated to the soil. The present study therefore aimed at determining the impact of wastewater irrigation on the habitat quality of water after soil passage and of soil after percolation by applying bioassays and chemical analysis. Lab-scale columns of four different soils encompassing standard European soil and three field soils of varying characteristics and pre-contamination were continuously percolated with treated wastewater to simulate long-term irrigation. Wastewater and its percolates were tested for immobilization of Daphnia magna and growth inhibition of green algae (Pseudokirchneriella subcapitata) and water lentils (Lemna minor). The observed phytotoxicity of the treated wastewater was mostly reduced by soil passage, but in some percolates also increased for green algae. Chemical analysis covering an extensive set of wastewater-born organic pollutants demonstrated that many of them were considerably reduced by soil passage, particularly through peaty soils. Taken together, these results indicated that wastewater-born phytotoxic substances may be removed by soil passage, while existing soil pollutants (e.g. metals) may leach and impair percolate quality. Soils with and without wastewater irrigation were tested for growth of plants (Avena sativa, Brassica napus) and soil bacteria (Arthrobacter globiformis) and reproduction of collembolans (Folsomia candida) and oligochaetes (Enchytraeus crypticus, Eisenia fetida). The habitat quality of the standard and two field soils appeared to be deteriorated by wastewater percolation for at least one organism (enchytraeids, plants or bacteria), while for two pre-contaminated field soils it also was improved (for plants and/or enchytraeids). Wastewater percolation did not seem to raise soil concentrations of classical organic pollutants and priority substances, while a significant retention was found for zinc and several organic micropollutants, particularly in the peaty soils, thus matching these soils' observed higher removal efficiency. Overall, our results demonstrate that benefits of wastewater irrigation can come with the cost of deteriorating soil habitat quality and depend on the respective soil and considered test organism. The approach employed here represents a feasible tool to assess these integrated effects at lab-scale while being predictive for scenarios at field-scale.

Imaging viscoelastic properties of live cells by AFM: power-law rheology on the nanoscale.

We developed force clamp force mapping (FCFM), an atomic force microscopy (AFM) technique for measuring the viscoelastic creep behavior of live cells with sub-micrometer spatial resolution. FCFM combines force-distance curves with an added force clamp phase during tip-sample contact. From the creep behavior measured during the force clamp phase, quantitative viscoelastic sample properties are extracted. We validate FCFM on soft polyacrylamide gels. We find that the creep behavior of living cells conforms to a power-law material model. By recording short (50-60 ms) force clamp measurements in rapid succession, we generate, for the first time, two-dimensional maps of power-law exponent and modulus scaling parameter. Although these maps reveal large spatial variations of both parameters across the cell surface, we obtain robust mean values from the several hundreds of measurements performed on each cell. Measurements on mouse embryonic fibroblasts show that the mean power-law exponents and the mean modulus scaling parameters differ greatly among individual cells, but both parameters are highly correlated: stiffer cells consistently show a smaller power-law exponent. This correlation allows us to distinguish between wild-type cells and cells that lack vinculin, a dominant protein of the focal adhesion complex, even though the mean values of viscoelastic properties between wildtype and knockout cells did not differ significantly. Therefore, FCFM spatially resolves viscoelastic sample properties and can uncover subtle mechanical signatures of proteins in living cells.

Search for over 2000 current and legacy micropollutants on a wastewater infiltration site with a UPLC-high resolution MS target screening method.

A target screening method using ultra high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS) was developed. The method was applied to 14 groundwater and 11 surface water samples of a former wastewater infiltration site, where raw wastewater was applied until 1985 and treated wastewater is applied since 2005. The measured data are compared with mass spectrometric data of over 2000 organic micropollutants (OMPs), including pharmaceuticals, personal care products, pesticides, industrial chemicals and metabolites of these classes. A total number of 151 and 159 OMPs were detected in groundwater and surface water, respectively, of which 12 have not been reported before in these matrices. Among these 12 compounds were 11 pharmaceuticals and one personal care product. The identity of 55 of the detected OMPs (35%) was verified by analysis of standard compounds. Based on the distribution in the study area, two groups of OMPs were clearly distinguished: current OMPs introduced with treated municipal wastewater since 2005 and legacy OMPs originating from infiltration of untreated wastewater until 1985. A third group included OMPs contained in historic as well as in current wastewater. During infiltration, OMPs with molecular mass >500 g/mol and log DOW > 3.9 were preferentially removed. Speciation had a strong impact with cationic OMPs showing high, neutral OMPs medium and anionic OMPs lowest elimination during infiltration. This target screening method proved useful to study a wide range of compounds, even in retrospect and at sites with poorly documented history and with a complex and variable hydrological situation.