Gold nanoparticles are being explored for in vivo applications, including imaging, drug delivery and therapeutic agents. It is now well-known that upon immersion in biological fluids, nanoparticles will be immediately overcoated by a "corona" of proteins, ions, and other species present in biological fluids. Understanding the basic thermodynamics, kinetics, conformation and orientation of bound biomolecules to nanoparticle surfaces is a critical piece of the larger picture of "what the cell sees." Using chemical tools, in this tallk I will discuss how to measure these processes, and how the nature of the surface has impact all the way to the level of cells and organisms.

There has been a surge of interest in applications of gold nanoparticles in biomedicine for imaging and therapeutic purposes in recent years. Thus understanding the behavior of gold particles inside polymeric or lipid carriers as well as the interaction of gold nanoparticles with lipid bilayers is highly desirable. While experimentally one can estimate the degree of encapsulation and stability of aggregates containing gold nanoparticles, the molecular details of interactions between nanocarrier and gold nanoparticles remains obscure. Computer modeling can provide valuable insights in this area. Using coarse-grained molecular dynamics simulations with a Martini force-field we investigate the interactions between hydrophobically-modified gold nanoparticles and a mixed lipid bilayer, nanodisk or vesicle. To obtain a lipid nanodisk a combination of at least two lipids with short and long tails is required. We investigate the equilibrium structure of self-assembled lipid with encapsulated single of multiple gold nanoparticles as well as probe the dynamic pathway for gold nanoparticle encapsulation. The effect of lipid composition (i.e. fraction of short to long lipids), gold nanoparticle size and length of hydrophobic tethers attached to gold nanoparticle will be discussed. Furthermore we will also discuss the interaction between encapsulated gold nanoparticles, which in some cases results in gold clustering. Cholesterol is one of the natural components of cell surface, thus it is of importance also to address the role of cholesterol in the behavior of self-assembled lipid structures and cholesterol-gold nanoparticle interactions. We will discuss and compare the behavior of gold nanoparticle containing lipid aggregates with and without cholesterol. The results of computer modeling will be compared with available experimental data and predictions will be made regarding the influence of the intrinsic properties of lipid aggregates and gold nanoparticle on the outcomes lipid/gold nanoparticle co-assembly and gold nanoparticles clustering inside the lipid aggregates.

The interaction of biomolecules (proteins, lipids et al) with synthetic nanoparticles topic of great interest in the allied fields of drug delivery, theranostics and bionanotechnology in general. The formation of the protein corona on any NP injected into the blood stream has been known for a while now, but methodologies to study the phenomenon are constantly being invented and reinvented, simply because there is no best method yet. This is a hurdle in the way of truly understanding the implications and adoption thereby, of nanomedicine.
Herein we demonstrate a robust protocol based on the Biorad XPR36 surface plasmon resonance setup to study affinities of serum proteins towards nanoparticles. We employ a hetero-bifunctional linker to immobilize our model nanoparticles (Au NPs) onto the surface of the chip. A layer of alginate matrix on the native gold surface of the chip keeps non-specific binding to a minimum. Thereafter, conditions were optimized for obtaining very stable immobilized AuNPs on the chip surface. Consequently, it became possible to study interactions between the proteins of interest, human serum albumin (HSA), fibrinogen, immunoglobulin G and apolipoprotein and the AuNPs. Through the use of this method, we could obtain kinetic data for these protein-NP complex formation interactions.
We also used the same method to demonstrate the significant retardation of corona formation on AuNPs functionalized with polyethyleneglycol (PEG) moieties.
With the identification of a suitable linker, the protocol can be used to study the interaction of any native or surface-functionalised NP with biomolecules. This can reveal important details as to what would be the final identity of a NP in the context of the physiological system. Instead of getting snapshots in time of the “state” of a NP incubated with serum proteins, this method lets the seeker have real time data - a possibility to visualize the formation of corona as and when it happens.

With the development of material sciences, the characterization of nanomaterials has become a critical issue in managing their fascinating size-dependent physical and chemical properties. Controlling these properties from the synthesis to the application phase, and consequently to their fate as a worldwide environmental and societal concern is becoming more and more imperative. The potential toxicity of nanoparticles needs to be evaluated when developing applications in a responsible way. Zinc oxide ZnO nanoparticles can be found largely as powders and dispersions with antibacterial, anti-corrosive, antifungal and UV filtering properties. ZnO nanoparticles can also be used for various applications ranging from food and cosmetics up to coatings agent and in the manufacturing of concrete. Research is actively being conducted towards in solar cells, photocatalysis, optical devices and sensors which have already started to show economic potential worldwide.
The principal techniques currently used to achieve the characterization of nanoparticles are physical and physico-chemical methods, such as Transmission Electron Microscopy (TEM), X-ray diffraction, and optical spectroscopies. All these analytical tools are excellent for global analyses of clusters and nanomaterials. Soft ionization mass spectrometry (MS) methods such as Matrix Assisted Laser Desorption Ionization coupled with Time of Flight MS (MALDI-TOFMS) have already proven their potential as tools in the nanometrology of small-sized II-VI quantum dots (QDs) such as CdS, CdSe, ZnS and ZnSe. Mass spectra of these nanocrystals are consistent with TEM and optical spectroscopy measurements [1-2].
In this paper, we present a joint physical/physico-chemical study and, more specifically, the first application of MALDI-TOF-MS to analyze small-sized ZnO QDs (3-3.5 nm diameter range) synthesized by sol-gel chemistry and stabilized through an aminosilane coating. The organic shell increases the QDs stability and dispersibility in aqueous solution. The ligands were first quantified by thermogravimetric analysis (TGA), then a careful investigation of the stability of ZnO QDs was initiated once these QDs were dispersed in different media (water, biological buffer,hellip;) for a period up to 6 weeks. Positive ion mode mass spectra showed a decrease in mass and consequently in diameter during aging, which can be ascribed to the degradation of ZnO QDs.
In conclusion, the unique combination of MALDI-TOF-MS and physico-chemical techniques brings new insights concerning the structure analysis, the stability and consequently the potential toxicity of ZnO QDs. This new strategy in nanometrology will be extended to other II-VI materials in the near future.
[1] A. Aboulaich, D. Billaud, M. Abyan, L. Balan, J.J. Gaumet, G. Medjahdi, J. Ghanbaja, R. Schneider ACS Appl. Mater. Interfaces, 4 (5) 2561-2569 (2012).
[2] M. Fregnaux, J.J. Gaumet, S. Dalmasso, J.P. Laurenti, R. Schneider Microelectron. Eng.108, 187-191 (2013).

Nanoparticle functionalization with peptides has proven to be a good strategy to inhibit particle aggregation, increase nanoparticle solubility and develop nanocarriers and biosensors [1,2]. Cell-penetrating peptides, such as, the transcription transactivation TAT peptide from human HIV-1 virus, have become a popular choice for transfection and other types of cellular delivery [3]. Studies have shown that TAT-peptide functionalized gold nanoparticles (AuNP) have the ability to penetrate cell membranes and localise in the nucleus [4,5], making them potentially a good system for drug-delivery applications, albeit with variable performance.
Detailed investigation of the effects of peptide concentration and distribution in the functionallayer at all-atom resolution can aid the design of gold nanomaterials for efficient drug delivery. With this in mind, we have performed an experimental and computational study on the effects of TAT peptide concentration on the structure and dynamics of the peptide layer on the 3 nm AuNP surface and its impact on the NP's membrane permeation capacity [6].
We show that cell internalisation of TAT functionalised nanoparticles is strongly dependent on TAT surface concentration and distribution. We present the mechanism by which TAT may become inactive upon surface immobilisation and demonstrate how the local environment of the functional peptide grafted onto the nanoparticle surface may affect properties essential for efficient cellular uptake. We found that a notably increased membrane activity correlates with the peptide backbone rigidity and increased basic residue solvent exposure attained at a particular concentration range of the functional peptides. The theoretically predicted membrane permeating capacity was confirmed experimentally while we also revealed molecular properties relevant to the experimentally observed inactivation with increasing TAT concentration. Our results suggest that nanoparticles may be engineered to specifically permeate membranes by targeted conjugation of peptides at specific concentration and surface distribution.
[1] P.D. Howes, R. Chandrawati, M.M. Stevens “Colloidal nanoparticles as advanced biological sensors” Science 346:1247390 (2014)
[2] A. Makarucha, N. Todorova, I. Yarovsky “Nanomaterials in biological environment: a review of computer modelling studies” Eur. Biophys. J, 40:103 (2011)
[3] N.A. Brooks, et al. “Cell-penetrating peptides: Application in vaccine delivery”, Biochem. Biophys. Acta, 1805:25 (2010)
[4] Z. Krpetic, et al. “Negotiation of intracellular membrane barriers by TAT-modified gold nanoparticles.” ACS Nano 5:5201 (2011)
[5] J.M de la Fuente, C.C. Berry “Tat peptide as an efficient molecule to translocate gold nanoparticles into the cell nucleus”, Bioconjugate Chem. 16:1176 (2005)
[6] N. Todorova, et al. “Surface presentation of functional peptides in solution determines cell internalization efficiency of TAT conjugated nanoparticles” Nano Lett. 14(9):5229 (2014)

Genetically Engineered Peptides for Inorganic solids (GEPI) are of a broad interest due to their capability in biofunctionalization of solids and utility as molecular linkers, erector sets and assemblers as well as tiny enzymes to synthesize inorganics under biologically viable conditions. Further refining combinatorial mutagenesis approaches, e.g., cell surface and phage display libraries, originally adapted from the principles of drug design, our laboratory has been experimentally selecting 1000s of GEPIs for a variety of noble metals (Au, Pt, Au, Ti), oxides (SiO₂, ZnO, Al₂O₃, ZrO₂), semiconductors (GaN), and minerals (HAp, quartz, calcite, aragonite, magnetite, diamond, sapphire, and graphite). To accelerate directed evolution processes, our Center also established bioinformatics methods to de novo design multifunctional chimeric constructs as second, third and higher generation peptides. Despite their short sequences (<25AAs) and, hence, intrinsically disordered structures in water, the versatility of GEPIs stem from their predictable folding conformations specific to a given surface with known physico-chemical characteristics. With the advent of 2D single atomic layer solids, we have recently developed rational approaches to design functional bio/nano interfaces via point and domain mutations. Based on the understanding of the fundamental surface phenomena, e.g., diffusion, assembly, and surface organization, these novel approaches allow us to intermolecular forces to construct peptide-enabled hybrid nanostructures with single atomic layer solids (graphene, MoS₂, WeSe₂, BN) allowing addressable chemical or physical functions. We will discuss latest developments in designing solid-binding peptides with specific surface recognition and assembly characteristics augmented by computational modeling (MD, MM, QM, kMC, etc.) and, finally, present examples in implementations in molecular-technology and -medicine, e.g., surface resistance spectroscopy, graphene-FET cancer biosensors, and nanophotonic devices. Funded by USA-ARO, NSF-DMR BioMat & MRSEC, NIH-NIDCR & NCI, JAPAN JST PRESTO.

Porous silicon (PSi) is a promising multifunctional biomaterial due to its large surface area for drug loading, tunable optics for biosensing, inherent biocompatibility and tailored biodegradation through control of surface chemistry. However, the promise of multifunctionality has been limited due to difficulties in chemical passivation for controlled degradation and relatively low sensitivity for biosensing in aqueous environments. Previously we showed how control over surface chemistry can significantly extend the lifetime of PSi materials, thus allowing the real-time monitoring of secretion from adherent cells in culture. In this presentation I will show how chemically modified mesoporous silicon photonic crystals can be used as “smart” substrates for monitoring cellular processes. First, I will show how loading the nanopores of PSi films with pharmacological compounds can be used for sustained release to guide the differentiation of mesenchymal stem cells to the bone cell lineage. Next, I will present a strategy to pattern gold islands in diverse geometric configurations across a PSi photonic crystal for use of orthogonal surface chemistries that differentially control the fate of adherent neural stem cells in culture. Finally, I will present the integration of a hydrogel defect layer into the photonic architecture as a means to detect secreted protease enzymes from adherent cells. Across all of these examples, careful control of the interface chemistry is critical to enabling the multifunctionality of nanostructured PSi materials for next generation implantable photonic devices and biomaterials for regenerative medicine.

Magnetic Nanoparticles are extensively used in theranostics: hyperthermia, enhancement of contrast in MRI, etc. The main problem with nanoparticles is their delivery to the target cells [1]. To this purpose, the overexpression of transferrin-receptor 1 in cancer cells makes transferrin a potential vehicle for nanoparticles [2]. Transferrin solubilizes Fe³⁺ in sera and when iron-loaded, it is recognized by receptor-1, which is anchored in the plasma membrane [3]. This Triggers the receptor-mediated endocytosis of the two proteins [4,5]. We develop here a "Trojan horse" system based on this endocytosis of the holotransferrine-receptor adduct to intracellular delivery of maghemite nanoparticles. Different sizes of maghemite (Fe₂O₃) superparamagnetic nanoparticles (5, 10 and 15 nm) were synthesized, coated with 3-aminopropyltriethoxysilane (APTES) and coupled to holotransferrin (TFe₂) by amide bonds. Each nanoparticle (NP) carried a known number of holotransferrins (TFe₂-NP). This transferrin construct was tested in vitro and remains active after grafting and interacts with its receptor rapidly (50 µs) with an overall dissociation constant (11 nM) [6]. HeLa cells were incubated for several time intervals with rhodamine-labeled TFe₂-NP and NPs. Confocal fluorescence microscopy showed that NPs do not cross the plasma membrane within 1 hour, whereas the constructs holotransferrine-maghemite are internalized in the cytosol in endosomes in less than 15 minutes (below). Furthermore, preliminary magnetophoresis results showed, in Lymphocyte T cells, that the rate of internalization of NPs grafted onto holotransferrine is three times larger than that of raw NPs in a culture media containing: 0, 10 and 55 % FCS. These very promising results seem to exclude the formation of a protein corona and validate our strategy. This hypothesis was also confirmed by molecular modeling. Thus, this nanohybrid system constitutes an interesting model for theranostic devices able to follow the main iron-acquisition pathway.
References
[1] A. Salvati, A.S. Pitek, M.P. Monopoli et al., Nat. Nanotechnol., 8 (2013) 137.
[2] T.R. Daniels, et al., Biochim. Biophys. Acta, 1820 (2012) 291.
[3] R.R. Crichton, Iron Metabolism: From Molecular Mechanisms to Clinical Consequences. Third Edition ed. West Sussex: J.Wiley & Sons (2009).
[4] A. Dautry-Varsat, A. Ciechanover, H.F. Lodish, Proc. Natl. Acad. Sci. USA, 80 (1983) 2258.
[5] M. Hemadi, P.H. Kahn, G. Miquel, et al., Biochemistry, 43 (2004) 1736.
[6] H. Piraux, J. Hai, P. Verbeke et al., Biochim. Biophys. Acta., 1830 (2013) 4254.

Although some peptides are known to bind to carbon-based solids, fundamental phenomena such as substrate specific peptide binding, conformational transformations and surface self-diffusion have not yet been fully understood. In light of the recent experimental observations of combinatorially-selected engineered solid-binding peptides that assemble to form long range ordered molecular nanostructures on atomically flat surfaces, e.g., graphite and MoS2, here we present a comprehensive simulation study, supported by experimental observations, on the structural and dynamic characteristics of engineered peptides using classical molecular dynamics (CMD) in aqueous environment. The focus of the current study is to determine adsorption free energy and diffusion dynamics of engineered dodecapeptides, including wild type (GrBP5-WT) and its 8 separate mutations with different anchoring domains and tail sequences. In particular, the specific role of the single residue mutations were studied which stabilize structural motifs of the peptide mutants on graphene and MoS₂ through molecular recognition of the surface lattice. Our calculations show that the strength of peptide adsorption on the solid surface can be regulated using even a single amino acid mutation, with levels ranging from no adsorption to moderate or strong binding concomitant with altered surface dynamics. A strong correlation can be developed between the rates of peptide self-diffusion on the solid vs their adsorption strength and molecular footprint that develops on the surface upon refolding. The specific role of self-diffusion is correlated with possible mechanism of self-assembly that result in long-range ordered biomolecular nanostructures on single layer materials which also include WSe2 and BN. Research supported by the NSF-MRSEC Program, DMR #0520567 and USA-DOE&’s Resources of NERSC via ERCAP with the Project ID. GrBP5, and NIDCR Fellowship Program.

Zinc oxide (ZnO) is a direct wide band gap semiconductor material (3.37 eV) useful for many electronic and optoelectronic applications including sensors, light emitting diodes, and photocatalytic devices. Bio-directed synthesis of ZnO is a promising technique for morphological control of nanostructures at benign temperature and pressure conditions. Specifically, ZnO synthesis utilizing free or unbound peptides in solution, as well as peptides chemically bound to planar substrates has been reported. Such peptides were able to initiate ZnO crystal growth and influence the relative growth rate of specific crystallographic planes, however they lacked the long range order needed for hierarchical assembly. In this work, one-dimensional (1-D) ZnO nanostructures were assembled using a M13 bacteriophage template that displayed a high copy number of well-ordered ZnO-binding peptides along its length. This ZnO-binding filamentous virus, which was approximately 6 nm in diameter and 880 nm in length, had a protein capsid primarily composed of thousands of a-helical major coat proteins, pVIII, arranged in a highly ordered five-fold symmetry. A ZnO-binding peptide was genetically displayed on each of these pVIII coat proteins creating a high aspect ratio scaffold with numerous densely-packed, highly-organized binding sites to aid in morphological control. The peptide fusion was discovered with combinatorial phage display using a pVIII library to ensure that its length, size, and chemical nature were compatible with the tightly packed nature of the pVIII major coat protein. The identified 8-mer ZnO-binding sequence was shorter in length and less positively charged than previously reported peptides with affinity for ZnO. Moreover, M13 bacteriophage that displayed this peptide on the pVIII protein coat were able to bind ZnO at densities nearly 100-fold more than the M13 wild-type virus. Transmission electron microscopy and scanning electron microscopy were used to characterize nanocrystal size, size distribution, morphology, and crystallinity of the assembled 1-D ZnO nanostructures. Furthermore, electron diffraction and energy dispersive x-ray spectroscopy were used for analysis of semiconductor crystal structure and composition. Two-terminal devices were fabricated from the 1-D ZnO nanostructures and photoresponse was measured. This simple synthesis method allows hierarchical assembly of 1-D nanostructures with specificity which can be translated to other materials systems with modification of the biological template.

Peptide-based strategies represent new avenues to access functional catalytic structures that operate under biological conditions. The activity of these structures arises from the unique biotic/abiotic interface developed between the peptides and the metallic nanocatalyst. In this regard, we have probed two different classes of biomimetic nanocatalytic materials where the peptide is either directly bound to the particle surface or used as a template to generate large metallic networks. Our results indicate that the localized interactions between the peptide and nanostructure play a key role in controlling the resultant catalytic functionality. These approaches can be applied for the generation of different monometallic materials and has recently been expanded for the fabrication of bimetallic alloys to enhance the catalytic reactivity. Furthermore, these materials can be used as model systems to probe fundamental reaction mechanisms, where our results support a metal leaching-based approach for C-C coupling reactions. By controlling the material morphology, biological interactions, and metallic composition, wide ranging catalytic functionality and potentially selective reactivity could be achieved using biomimetic materials that operate under green conditions.

The evolution-optimized processes of biomineralization yield multifunctional biominerals consisting of bioorganic and inorganic components. The properties of these nanostructured solids are often superior to their inorganic counterparts. Biomineral formation occurs in aqueous solution at ambient conditions and involves biopolymeric templates and interfaces that govern the mineralization of the inorganic components. The generation and the structure of these templates are genetically determined. Accordingly, the genetically controlled structuring of biotemplates represents also a promising approach for the design of non-biogenic inorganic functional materials applicable in functional devices.
Recently, we successfully employed the wildtype of the tobacco mosaic virus (TMV) as a protein-based biotemplate for the mineralization of the inorganic functional material ZnO. In this case, a reaction solution was used, yielding ZnO particles with a size in the range of 5 nm. The nanoparticles were selectively self-assembled on the virus surface [1].
Here, we present how the biotemplate formation by self-assembly of TMVs can be triggered by the genetic modification of the virus. For that purpose, monolayers made of the wildtype and several TMV mutants were prepared applying a convective assembly approach. The impact of the genetic modification on the self-assembly performance was studied. Pronounced differences in the surface arrangement of the different TMV variants were observed. The obtained TMV structures with different packing density were mineralized with ZnO. The mineralized ZnO structures directly display the morphologies of the biotemplates. Subsequently, the obtained TMV/ZnO materials were investigated with respect to their functional properties. The semiconducting behavior was studied by the integration of these bio/inorganic hybrids into field effect transistors. The results reveal that the transistor behavior can be modified and tuned by the genetic modification of the virus-based biotemplates. In addition, nacre-like multilayers made of the TMV and ZnO were prepared. The materials show an improved mechanical performance compared to the pure inorganic material due to the presence of the introduced bio/inorganic interfaces.
[1] P. Atanasova, D. Rothenstein,J. J. Schneider, R. C. Hoffmann, S. Dilfer ,S. Eiben, C. Wege, H. Jeske, J. Bill, Adv. Mater. 23 (2011) 4918

Material formation in nature is precisely controlled in all aspects from crystal nucleation, growth to assembly to deliver superior functions. Specific biomolecule-material interactions have been hypothesized to play important roles in these processes. Proteins, polymers and small molecules have been extensively explored to replicate the degree of control in material formation in vitro and for nonbiogenic materials. However the organic-inorganic interfacial interaction is still far from being understood which hinders the further advancement of biomimetic material formation. In this talk I will share our efforts in decoding the myth of biomolecular specificity to material surface and their roles in controlling crystal nucleation and growth. The selection of facet specific short peptides and their abilities in guiding predictable morphology control of Pt nanocrystals will be first demonstrated. Then detailed experimental and theoretical studies on binding mechanism will be discussed. At the end of the talk, the discovered molecular signature for facet specific adsorption will be applied to design small molecules which can modulate the nucleation/growth of the Pt nanocrystals to deliver the expected nanostructures. These studies open up opportunities in understanding the molecular details of inorganic-organic interface interaction, which can one day lead to the development of a library of molecular functions for biomimetic materials design and engineering.

Nature provides the inspiration for engineering structural- and processing-design criteria for materials. With a growing understanding of the molecular processes involved, bio-inspired strategies are increasingly explored to develop the next generation of biomaterials. There are several challenges in these strategies including replicating the hierarchical organization of biological materials, organization that provides multi-scale structure/property interdependence. The interfacial interactions become critical in tuning the individual components towards the changing functional needs. There is a need for strategies that can control self-organization at a molecular level and thus provide predictability over the biological and inorganic interfaces. In biological systems, proteins conduct various functions through their unique molecular recognition, self-assembly and templating properties. In addition to their role in biomineralization, proteins perform a wide spectrum of additional functions ranging from catalysis to self-repair. The approach includes decoding the peptide-material interactions, and using these foundations to develop self-organized and multifunctional hybrid systems. Genetic engineering and biochemical conjugation are used to produce chimeric peptides and fusion proteins that can controllably bind to a given solid including variety of materials. Here we demonstrate how we utilize the engineered peptides based multifunctional constructs for biological mineralization by developing bio-enabled routes bridging cell programmability to materials as well as biological self-assembly based surface functionalization while reserving the biological activity intact. Our results showed that the peptides can be tailored as major enabling biomolecules in the formation of mineral microlayers on implantable materials with a potential for dental and craniofacial restoration. We also demonstrated that peptide based approach can be applicable to control bioactivity and prevent infections at the implant-host interface. The peptide coatings can be tailored to carry various active entities as targeted delivery constructs. The efficacy of the different sets of multi-functional peptides are demonstrated for various implant materials.These methods will be discussed by introducing orientation and spatial distribution control towards applications in advanced materials fabrication and restorative and regenerative medicine . Multifaceted approach developed provides bio-enabled, single step processes as a key route in simplifying the advanced materials fabrications in biologically friendly conditions. We greatly acknowledge the funding from NIHR-NIAMS, NSF and KU Internal Funds.
Zhou Y, M.L. Snead, C. Tamerler, Nanomedicin nanotechnology, Biology and Medicine, 2014, in press
Yazici H, Fong H, Wilson B, Oren EE, Amos F, Evans JS, Zhang H, Snead ML, Sarikaya M, Tamerler C, ACTA Biomaterilia, 2013; 9(2): 5341-5352

The fluorescent tracer, PDMPO (2-(4-pyridyl)-5-((4-(2 dimethylaminoethylaminocarbamoyl) methoxy)phenyl) oxazole), has unique silica specific fluorescence leading to its use in biology to understand the processes underpinning biosilicification. However, the mechanism by which silica specific fluorescence is produced is not well understood nor is the response to local environmental variables like solvent and pH. To understand the silica specific behaviour, we investigated the spectroscopic properties of PDMPO in a range of environments: pH, solvent, and in the presence of silica forming compounds (tetraethoxysilane (TEOS)) and silica particles (23-180 nm diameter). The spectroscopy was supported with dynamic light scattering and zeta potential measurements to improve our understanding of the dye-silica interaction.
This presentation will describe the results of our investigation.

Mimicking the structure and morphology of biominerals, such as sea urchin spines, bones or teeth, under laboratory conditions is a challenging task, where the control over the nucleation and growth of crystalline phases is often achieved by utilizing soluble additives present within the reaction mixture.¹ Recent results suggest that the compartmentalized volumes available for mineralization processes within living cells or tissues also contribute to the final structure and morphology of the produced materials. Experiments with calcium carbonate crystals grown within polycarbonate track-etched membrane pores revealed that the confined volume had a significant influence on the crystallization process and resulted in the formation of single-crystalline calcite or vaterite rods, which were characterised by high aspect ratios.^2,3 A similar methodology was adopted to grow polycrystalline hydroxyapatite rods where the individual particles showed a marked preferential orientation.⁴
In this work we investigated the possibility of applying this bio-inspired synthetic route to precipitate hydroxyapatite crystals within track-etched membrane pores in the presence of soluble additives. As this polymorph of calcium phosphate is an important component of human bones and teeth, our intention was to understand the principles governing its formation. The utilized reaction conditions resembled that of the human body (pH, temperature), but the mineralization process took place within a confined volume that mimicked the compartmentalized environment of collagen fibrils.
TEM imaging of the obtained material revealed that the product particles exhibited a rod-like morphology and were composed of multiple hydroxyapatite crystals. The length of the rods was ca. 5 mu;m and their diameters were consistent with the diameters of the membrane pores. Electron diffraction analysis allowed us to study the crystallographic orientation of the rods. We observed that membranes with small diameters (25 nm - 50 nm) directed the orientation of the hydroxyapatite plates within the rods and that further reduction of the pore size to 10 nm resulted in the formation of rods that were indistinguishable from single crystals. In subsequent experiments, habit modifiers, such as amino acids, were added to the reaction mixture and their influence on the structure and integrity of the obtained rods was investigated. We learnt that by using glycine or glutamine it was possible to produce single-crystalline rods even in membranes with larger pore diameters. These results provided us with unique insight about how nature may control the deposition of inorganic phases within the human body.
[1] Meldrum, F. & Cölfen, H. (2008), Chem. Rev., 108(11), 4332-4432.
[2] Kim, Y.-Y. et al. (2011), Angew. Chem. Int. Ed., 50(52), 12572-12577.
[3] Schenk, A. S. et al. (2014), Chem. Comm., 50(36), 4729-4732.
[4] Cantaert, B., Beniash, E. & Meldrum, F. C. (2013), Chem. Eur. J., 19, 14918- 14924.

Biomimetic approaches utilize biomolecules and synthetic analogs to produce materials of controlled chemistry and function under conditions compatible with biological processes. Utilizing biomimetic synthesis, titania materials were generated under the influence of biomolecules and polymers (e.g. polyethyleneimine (PEI), protamine, and lysozyme) from aqueous Ti-containing precursors. These biomimetically synthesized products are hybrid materials that contain the mediating biomolecule as well as TiO₂. Unfortunately, biomimetic techniques typically produce amorphous or poorly-crystalline titania materials or require specialized, recombinantly produced proteins to produce highly-crystalline rutile or anatase TiO₂. In this study, we have investigated the devitrification of biomimetically synthesized titania materials under hydrothermal conditions that are similar to natural microbial environments. This processing route yielded bio-hybrid /TiO₂ nanomaterials that displayed photocatalytic activity exceeding that of commercially available titania photocatalyst (Aeroxide P25). Furthermore, the production of highly-crystalline titania under biomimetic conditions is advantageous, as the organic component of the bio-hybrid material is preserved and available for further functionalization. Indeed, the chlorination of these organic constituents yields halamine compounds that provide potent antimicrobial activity to the material system, independent of illumination conditions.

The hybridization of free oligonucleotides to densely-packed, surface-bound arrays of DNA is perturbed considerably compared to solution-phase binding of identical strands. Whereas experimental information from the DNA microarray literature has revealed qualitatively that hybridization is diminished in this context, acquisition of detailed thermodynamic data has proved challenging, as individual binding events non-linearly influence subsequent binding events. In this work, we utilize spherical nucleic acid (SNA) gold nanoparticle conjugates, i.e. gold nanoparticles densely functionalized with oligonucleotides, as a platform to investigate these effects in a quantitative manner. Because SNAs are colloidal objects with a well-defined concentration and DNA density, extraction of relevant thermodynamic binding data is more straightforward than when oligonucleotides hybridize to ill-defined surface monolayers of DNA. SNAs are observed to hybridize complementary strands with negative cooperativity, i.e. each binding event destabilizes successive binding events. DNA hybridization is therefore an ever-changing function of the number of strands already hybridized to the particle. Thermodynamic quantification of this behavior reveals a three order of magnitude decrease in the binding constant for the capture of a free oligonucleotide by an SNA conjugate as the fraction of pre-hybridized strands increases from 0 to ~30%. Increasing the number of pre-hybridized strands: (1) imparts an increasing enthalpic penalty to hybridization as the SNA accumulates charge that makes binding more difficult, while simultaneously (2) decreasing the entropic penalty to hybridization by inducing outstretched conformations in the remaining single strands which makes binding more favorable. Therefore, hybridization of free DNA to an SNA is governed by both electrostatic effects related to incomplete charge neutralization, and an effect consistent with allostery where hybridization at certain sites on an SNA modifies the binding affinity at a distal site through conformational changes to the structure. Leveraging these insights allows for the design of conjugates that hybridize free strands with significantly higher efficiencies, some of which approach 100%. This analysis provides important fundamental insight into the enthalpic and entropic consequences of packing charged DNA strands into a dense monolayer and, as a result, contributes new design considerations for high-density DNA microarrays and nanoparticle-based detection and therapeutic platforms.

DNA-mediated assembly strategies that take advantage of rigid building blocks, functionalized with oriented oligonucleotides to create entities with well-defined “valencies”, have emerged as powerful new ways for programming the formation of crystalline materials. With such methods, one can make architectures with well-defined lattice parameters, symmetries, and compositions, but to date they have been confined primarily to the use of hard inorganic nanoparticles or highly branched pure nucleic acid materials. In contrast, Nature&’s most powerful and versatile nanostructured building blocks are proteins, and are used to effect the vast majority of processes in living systems. In addition, unlike most inorganic nanoparticle systems, proteins can be made in pure and perfectly monodisperse form, making them ideal synthons for supramolecular assemblies. However, the ability to engineer the crystallization of proteins is not straightforward, and methods for deliberately making superlattices from them with both long-range order and predictable lattice symmetries and constants do not exist. To date, the primary methods for making protein lattices have relied on the use of natural protein-protein interactions, interactions between proteins and ligands on the surfaces of inorganic nanopaticle (NPs), metal coordination chemistry, small molecule ligand-protein interactions, or genetically fusing protein complexes with specific symmetries. Here, we introduce a new method for effecting protein crystallization by trading protein-protein interactions for complementary oligonucleotide-oligonucleotide interactions. By using different proteins functionalized with the appropriate oligonucleotides, along with the design rules introduced for inorganic systems, we show that both different combinations of proteins, and proteins and inorganic nanoparticles, can be assembled deliberately into preconceived lattices and, in some cases, well-defined crystal habits. Importantly, in the case of the two enzymes studied, activity is maintained in the superlattice states. This work is a convincing demonstration for how DNA can be used as a means for assembling many readily accessible functional proteins into ordered materials, regardless of the atomic composition of the proteins.

The complexity of the molecular recognition and assembly of bioticminus;abiotic interfaces on a scale of 1 to 1000 nm can be understood more effectively using simulation tools along with laboratory instrumentation. Current capabilities and limitations of atomistic force fields are discussed and a strategy to obtain dependable parameters for inorganic compounds is explained that has been extensively tested. Resulting parameters for silicates, aluminates, metals, oxides, sulfates, apatites, and other compounds are summarized in the INTERFACE force field. The INTERFACE force field operates as an extension of common harmonic force fields (PCFF, COMPASS, CHARMM, AMBER, GROMACS, and OPLS-AA) by employing the same functional form and combination rules, enabling simulations of an unlimited number of inorganicminus;organic and inorganicminus;biomolecular interfaces. The parameterization builds on an in-depth understanding of physical chemical properties on the atomic scale to assign each parameter, especially atomic charges and van der Waals constants, as well as on the validation of surface properties for each compound in comparison to measurements. The approach eliminates large discrepancies between computed and measured bulk and surface properties of up to two orders of magnitude using other parameterization protocols and increases the transferability of the parameters. As a result, a wide range of properties can be computed in quantitative agreement with experiment, including densities, surface energies, solidminus;water interface tensions, anisotropies of interfacial energies of different crystal facets, adsorption free energies of biomolecules on nanocrystals, thermal and mechanical properties.
Examples of biomolecule recognition on nanostructured metals, silica, and apatites illustrate insight into molecular recognition in 3D atomic resolution towards improving catalysts, sensors, and therapeutics. The adsorption mechanism of peptides on metal surfaces was discovered computationally to involve coordination of polarizable atoms in the peptide (C, N, O) with epitaxial sites on the metal surface, as well as contributions from induced charges. Computation predicts facet-specificity and preferred nanoparticle shape in correlation with laboratory synthesis, X-ray, TEM, and spectroscopy, as well as catalytic reactivity using reactive extensions of the INTERFACE force field. For oxidic minerals such as silica, the explicit consideration of the surface chemistry as a function of pH and particle size allows quantitative predictions of peptide adsorption, including sensitive point mutations. Similarly, the nanostructure of hydroxyapatite and its major facets varies strongly with pH, showing very different surface definition and binding mechanisms of proteins and osteoporosis drugs. A wealth of information in atomic resolution is becoming accessible by such computer-aided models in partnership with synthesis and characterization efforts to accelerate materials discovery.

Beyond its primary role as the carrier of genetic information, DNA has rapidly become one of the most important molecules in nanoscience, with its unparalleled programming and structural characteristics it has allowed the development of many novel nanoscale systems. RNA has also recently attracted significant interest for a variety of applications: the sequences found in vivo provide direct information about gene expression, its role in cell regulation has become evident, and the fact that it is naturally amplified inside the cell makes it an ideal target for sensor applications. For all of these reasons it is important to understand the electronic properties of both DNA and RNA duplexes. The electronic properties of DNA have been the subject of intense research over the last several decades, and incredible progress has been made using photochemical and electrochemical measurements to understand DNA&’s charge transfer characteristics. However, direct DNA conductance measurements have resulted in conductance values that span many orders of magnitude. Here we report on single-molecule conductance measurements performed on DNA duplexes and DNA:RNA hybrids using the STM-break junction approach in order to obtain the large datasets necessary for acquiring reproducible conductance values. With these measurements we demonstrate the necessity of controlling the local environment of DNA in order to be able to obtain reproducible conductance values and understand the charge transport mechanisms. Moreover, we extend this capability to study the charge transport characteristics of RNA:DNA hybrids, and show that the alternative structures in these duplexes result in significantly different transport properties. We will also discuss the effects of sequence changes, such as the roles of A:T, A:U, and G:C base pairs within the stack on the transport characteristics as well as the length dependence of the conductance. These measurements are coupled with electronic structure calculations and Green&’s function based transport calculations to understand the underlying differences between the various duplexes and sequences. Finally, we will examine the utility of single-molecule conductance measurements for identifying biologically relevant sequences.

The adsorption of biomolecules is of relevance to their purification, the response of the body to implants, including their biocompatibility, fate of nanomaterials in vivo, including their toxicology, and protein-mediated self-assembly of nano-structured materials and systems. This wide relevance has motivated us to undertake a program of research focused on using molecular simulation to build fundamental understanding of adsorption and transport of proteins on various surfaces, including metal, graphite, graphene and self-assembled monolayers, and to exploit this new understanding to develop technologies. In this talk, I will outline the progress of this work, including: (1) an until now unavailable molecular-level understanding of protein adsorption at various classes of fluid/solid interface [1]; (2) an efficient means of predicting free energy of adsorption of peptides at liquid/solid interfaces [2] and more rapid means of ranking adsorption propensity of peptides; (3) work focused on the molecular simulation-based design of peptides for self-assembly of graphene-based nanomaterials that have been realised in the lab [3]; and (4) the elucidation of the adsorbed conformations of MP-11, a microperoxidase, on graphene [4] that is relevant to a range of applications, including biofuel cells, bioresponsive logic circuits and biosensors amongst other things.
M. J. Penna, M. Mijajlovic, and M. J. Biggs, J. Am. Chem. Soc., 136, 5323 (2014). DOI: 10.1021/ja411796e.
M. Mijajlovic, M. J. Penna, and M. J. Biggs, Langmuir, 29, 2919 (2013). DOI: 10.1021/la3047966.
M. J. Biggs, M. V. Kiamahalleh, M. Mijajlovic and M. J. Penna, Compositions comprising self assembled carbon based structures and related methods, Provisional Australian Patent Application AU2014900273 (2014). http://www.ipaustralia.com.au/applicant/adelaide-research-and-innovation-pty-ltd/patents/AU2014900273/
M. Mijajlovic, Y. Sun, M. J. Penna, M. V. Kiamahalleh, W. Yang and M. J. Biggs, submitted (2014).
The supercomputing resources for this work were provided by eResearchSA and both the NCI National Facility at the Australian National University and the iVEC Facility at Murdoch University under the National Merit Allocation Scheme. The support of the Australian Research Council (DP130101714) is also acknowledged.

Cell-material interaction increase is a fundamental requirement in tissue engineering and in several biomedical applications. In this context, protein adsorption has been largely used to decorate the biomaterial surfaces without any preliminary surface modification as an alternative to chemical methods and covalent modification. On the other hand, protein adsorption deals with unpredictable, nonspecific and unstable interactions with the cells and the material surfaces.
As a possible route to overcome these disadvantages, we proposed to use amino-acid sequences composed by two domains: a part combinatorially selected to bind to a given material and a bioactive part able to stimulate cell-adhesion¹. Toward the applicability of this method it is fundamental to understand if the parts composing the two domains are still able to work together as bi-functional linkers, preserving a stable and specific interaction toward the biomaterial surface and keeping, at the same time, a suitable signal molecule display.
Along this way we have studied the interaction between bi-funcional peptides and gold surface employing classical molecular dynamics with specific parameters² able to describe the interaction between biomolecules and inorganics.
We have highlighted both the enthalpic and the entropic contributions in the adsorption event. As a case study we have examined a gold-binding peptide³ selected via combinatorial methods and extensively experimentally characterized, named AuPhi;3, alone and in the context of longer sequences together with the RGD and IKVAV cell binding motifs. The simulation results showed appreciable differences among the sequences in terms of the number of adsorbed residues and the conformational behavior at the gold interface. The case of the IKVAV-AuPhi;3 sequence as a bi-functional peptide results in a good possibility to employ non covalent binding moieties to promote surface bio-activation of gold.
1 Hersel, U.; Dahmen, C.; Kessler, H. RGD modified polymers: biomaterials for stimulated cell adhesion and beyond. Biomaterials 2003, 24, 4385-4415.
2 Heinz, H.; Vaia, R.; Farmer, B.; Naik, R. Accurate simulation of surfaces and interfaces of face-centered cubic metals using 12- 6 and 9- 6 Lennard-Jones Potentials. The Journal of Physical Chemistry C 2008, 112, 17281-17290
3 Causa, F.; Della Moglie, R.; Iaccino, E.; Mimmi, F.; Marasco, D.; Scognamiglio, P. L.; Battista, E.; Palmieri, C.; Cosenza, C.; Sanguigno, L.; Quinto, I.; Scala, G.; Netti, P. Evolutionary screening and adsorption behavior of engineered {M13} bacteriophage and derived dodecapeptide for selective decoration of gold interfaces. Journal of Colloid and Interface Science 2013, 389, 220-229.

Material binding peptides are proving to have great potential in improving material synthesis and advancing device fabrication, however, their specificity and interaction mechanisms with target surfaces remain largely elusive. This study contributes to the developing understanding of fundamental principles through which ZnO binding peptides (ZnO-BPs) interact with and modify ZnO growth/morphology. ZnO-BPs used were the reported phage display (PD) identified sequence (G-12 (GLHVMHKVAPPR) and its derivative, GT-16 (GLHVMHKVAPPR-GGGC)) as well as novel sequences generated from post selection modifications including alanine mutants of G-12 (G-12A6, G-12A11, G-12A12) chosen on the basis of peptide stability calculated in silico. Two approaches were used to study interaction of ZnO-BPs with ZnO. Firstly ZnO growth was monitored in the absence and presence of ZnO-BPs during solution synthesis using two different growth routes; the Zn(NO₃)₂middot;6H₂O-HMTA system and the ZnAc₂-NH₃ system. Secondly, isothermal titration calorimetry (ITC) was used to characterize thermodynamic changes during interaction of ZnO with ZnO-BPs. The outcomes of the ZnO synthesis studies demonstrated that a single ZnO-BP can adopt different sequence and concentration dependent mechanisms to control ZnO growth/morphology. The specific synthesis system used dictated the products which ZnO-BPs could interact with and consequently modify crystal nucleation and growth processes. One of the outcomes of the study is the demonstration of the role of histidine within ZnO-BPs in interaction with ZnO and stabilization of LBZs. Analysis of the thermodynamics of ZnO-BP-ZnO crystal interactions using ITC yielded isotherms comprising both an endothermic and an exothermic event. Measured #8710;G values were between -6 and -8.5 kcal/mol and high adsorption affinity values indicated the occurrence of favourable ZnO-BP-ZnO interactions. Predictive control of material formation processes by peptides can be achieved through a clear understanding of the growth processes and interaction mechanisms.

Peptide-enabled synthesis of inorganic nanomaterials is attractive due to the potential of creating materials with optimized/tunable properties through sequence manipulation. While there is a large amount of literature available on the fabrication of nanoparticles using peptides, comparatively little is known the nanomaterial&’s in-depth atomic structure. Such information is an important prerequisite for rational sequence design which can result in nanomaterials with tunable properties. Using a hybrid experimental/computational approach, sequence-dependent structure/function relationships are assessed and predicted by combining X-ray characterization methods and molecular dynamics simulations for monometallic and bimetallic systems. First, extended X-ray absorption fine-structure spectroscopy (EXAFS) is performed on peptide-capped nanoparticles to determine sequence-dependence in local chemistry and structural disorder. From there, high-energy X-ray diffraction (HE-XRD) data was taken and coupled with atomic pair distribution function (PDF) analysis to obtain sub-atomic scale structural details over the entire nanoparticle structure. Reverse Monte Carlo simulations of the PDF data, which are guided by the EXAFS data and other constraints, are used to obtained nanoparticle configurations which demonstrate sequence-dependent structural disorder. From there, molecular dynamics methods are used to relax particle structures, model peptide morphology on the experimentally-determined structures, and finally used to assess catalytic properties to provide sorely lacking sequence-dependent structure/function relationships. The aforementioned methodology presents a widely-applicable route for peptide-enabled systems to provide sequences design rules toward the ultimate goal of rational nanomaterials design.

Peptides are versatile biomolecules that play a strong role in physiological processes through their role as signalling molecules. Peptide coatings are therefore being utilized to modify how the body recognizes and responds to a variety of implants, including those utilized in neural interfaces. In this work, we present a strategy for attaching peptides (derived from the neural cell adhesion molecule) to the surface of fine (50 µm) polyimide-insulated microwires, towards modifying response of astrocytes and microglia (the cells involved in the foreign body response in the central nervous system) to the microwires. The peptide modification process includes oxygen plasma treatment of the polyimide surface, covalent attachment of a silane linker ((3-Aminopropyl)triethoxy silane (APTES)) to this plasma treated surface and covalent attachment of the peptide to the silane layer through 1-Ethyl-3-(3-dimethylaminopropyl)carboiimide (EDC) chemistry. The density of the silane layer deposited under different pH conditions (pH 4 and 6) was determined using XPS, and it was found that deposition at pH 6 yielded multilayers of adsorbed silane while deposition at pH 4 resulted in the formation of monolayers. The stability of the silane was examined under physiological conditions: adsorbed APTES multilayers degraded quickly under physiological conditions whereas monolayers formed at pH 4 were stable under the same conditions. The density and uniformity of surface coverage of the peptide achieved under different reaction conditions (peptide concentration, time, temperature, EDC concentration) was analyzed through imaging by fluorescent microscopy of a dye (5 Carboxytetramethylrhodomide (TAMRA)) bound to the peptide. The cellular response was observed by inserting wires functionalized with an adhesion peptide and un-modified reference wires into 3D gels seeded with primary cultured astrocytes and microglia; increased astrocyte density was observed at the surface of the functionalized wires as compared with reference wires. Such coatings could play a role in mediating the foreign body response to implants in the central nervous system. Our methods could also be utilized in future applications requiring the attachment of diverse peptides to polymers.

In general, an engineered virus for binding a target material discovered via a high-throughput evolutionary screening process possesses a limited recognition or nucleation function for the selected material. In this study, M13 bacteriophage (phage) was engineered for the use as a versatile template for preparing various nanostructured materials via genetic engineering coupled to enzymatic chemical conversions. First, we engineered the M13 phage to display TyrGluGluGlu (YEEE) on the pVIII coat protein and then enzymatically converted the Tyr residue to 3,4-dihydroxyl-L-phenylalanine (DOPA). The DOPA-displayed M13 phage were able to perform two functions: assembly and nucleation. The engineered phage assembled various noble metals, metal oxides, and semiconducting nanoparticles into one-dimensional arrays. Furthermore, the DOPA-displayed phage triggered the nucleation and growth of gold, silver, platinum, bimetallic cobalt-platinum, and bimetallic iron-platinum nanowires. This versatile phage template enables rapid preparation of phage-based prototype devices by eliminating the screening process, thus reducing effort and time.

Gold nanoparticles have been widely explored in biomedical applications as active engineered materials for diagnosis and therapy. With the great potential of using these nanocomposites in biological systems, such as in the manufacture of small materials for biotechnological application, it is very interesting to understand with details how these materials interact at the molecular level with cell membranes in living systems. In this study we investigated the interactions between gold nanorods and cell membrane models using the technique of vibrational sum frequency generation-SFG spectroscopy, specific for interfaces. The membrane models comprised Langmuir phospholipid monolayers composed by DPPC (dipalmitoylphosphatidylcholine) and DPPG (dipalmitoylphosphatidylglycerol). The interaction between the gold nanorods and the membrane models was revealed via SFG as the decrease in the CH₃ band at 2880 cm^-1 in comparison to the band at 2942 cm^-1. The investigation of the interactions between nanomaterials and membranes using SFG may be relevant for nanotoxicological studies.

Tailings are the byproducts left over from mining and extracting resources, such as bitumen from the oil sands or minerals from ores. The substances found in tailings ponds depend on the type of mining operation. However, many of them are very precious materials, such as noble metals, semiconductors, rare earth elements, etc.. An engineered new material, which is environmentally sustainable and poses the ability to effectively mine the precious materials from the tailings, is in demand.
Here we present our work on preparing novel biomaterials composed of gold binding peptides conjugated on the surface of magnetic nanoparticles (AuBP-MNPs) for gold mining. These biomaterials were designed to possess two significant functions: the surface conjugated gold-binding peptide, obtained from cell surface display, will recognize and selectively bind to gold, while the magnetic nano-sized core will response and migrate corresponding to the applied external magnetic field. This will allow the smart biomaterials to mine an individual material (gold) from a pool of mixture without excessive solvent extraction, filtration and concentration steps.
The AuBP-MNPs were prepared using a three-step protocol under aqueous reaction conditions at room temperature. Highly uniform quasi-spherical iron oxide magnetic nanoparticles (~70nm) were synthesized by using hydrothermal reaction with slight modification. Then amine or carboxylate groups were introduced onto the surface of magnetic nanoparticle. Finally, gold binding peptides (AuBP1 and AuBP2) were conjugated to the functionalized surface via coupling reaction. Structure, morphology, and magnetism of the AuBP-MNP were thoroughly investigated by means of FTIR, SEM, HR-TEM, SAED, XPS, Zeta-potential, TGA, and vibrating sample magnetometer. Surface coverage of the peptides was quantified by UV-Vis spectroscopy. Applying AuBP-MNPs to Au nanoparticle solution (1-3 nm) gave a dramatic reduction of Au nanoparticles concentration via magnetic separation.
In the next step, the pH, ionic strength, and temperature will be studied extensively in order to identify a possible condition for biomaterial-gold dissociation. We are aiming to make these biomaterials recyclable and able to work in a continuous flow process.

In this work, an amorphous Fe rich amorphous structure designed based on competitive atomic cluster model was synthesized and characterized successfully. The constituent zero-valent iron (ZVI) has excellent activity and efficiency for decolourization of Orange G (OG) solution at room temperature. The decolourization is characterized by UV-Vis spectrum and pseudo first-order kinetics. The X-Ray Micro fluorescence spectrometer, Inductively Coupled Plasma Optical Emission Spectrometry and Scanning Electron Microscope were employed to trace the ZVI. The consumption of ZVI to destabilizes the local atomic arrangement and yields the phase separation of Fe at surface, responds to the high activity and the catalysis for decolourization. This observation is in accordance with the change of k₁ 0.011min^-1 to k₂ 0.047min^-1, which is supported by the cyclic decolourization test. This work provides a new strategy to design multi-functional metal materials and indicates their brilliant future in practical application.

Reverse osmosis is one of the most mature and attractive technologies for water purification, owing to the selective nature of semi-permeable membranes. Conventional polymer membranes have a structure of three layers: a polyester web support, a micro-porous interlayer (ie. Polysulfone) and an ultra-thin selective layer made of aromatic polyamide via interfacial polymerization. In order to achieve high salt rejection, the membrane pore size is usually less than 1 nm. As a result the water flux is extremely low and high pressure is required during the separation process. In order to enhance water permeability and reduce cost, AquaporinZ (AqpZ), a highly selective water channel protein from E. coli, can be introduced to form an efficient biomimetic membrane to overcome these challenges.
Though AqpZ-embedded membranes mediated by proteopolymersome or planar lipid bilayers have been reported, they still have several shortcomings: 1) defects due to the materials and fabrication techniques result in contaminants passing through the membrane; 2) Planar membranes containing AqpZ were not able to cover porous substrates effectively; 3) the selective layer and the substrate may delaminate due to the weak interaction; 4) fabrication methodologies are not scalable.
In order to solve the aforementioned problems of AqpZ-membranes, we use a novel synthesis and fabrication strategy. Rather than lipid or block co-polymers, we utilize a short amphiphilic peptide to protect and improve the stability of AqpZ in the membrane. This amphiphilic peptide has a sequence of alternating polar and nonpolar residues to form beta-sheet in solution, and in the presence of protein it wraps around to form a protective beta-barrel. We then functionalized the peptide with an end group that can react with the porous membrane substrate. As a consequence, we are able to directly control the assembly of protein on the substrate. We chose polysulfone as our starting material for membrane casting because of its high chemical compatibility and durability. We first functionalized polysulfone with azide group, then we casted the membrane using a liquid-induced phase inversion technique. Pore developing agent is added during the casting process to form membranes with pores of controlled size. The azide functionality around the pores provides the provision for specific attachment of functional proteins. Thus, the AqpZ-peptide complex can be covalently cross-linked to the polysulfone substrate through mild click chemistry resulting in a robust and scalable protein-loaded biomimetic RO membrane. The stability and desalination performance of synthesized membranes are currently under investigation.

In the context of food safety, contamination of food-contact surfaces with pathogenic bacteria is a global concern. This work investigates the potential of hydrophobically-modified, nanoporous silica aerogel as a bacterial anti-adhesion food-contact surface. The bacterial anti-adhesion efficacy of the silica aerogel, hydrophilic nonporous silica (negative control with no bacterial anti-adhesion property), and hydrophobically-modified nonporous silica (positive control with some bacterial anti-adhesion property) were evaluated using dip inoculation with Salmonella Typhimurium LT2 and Listeria innocua NADC 2841 at 8.8 to 9.1 log CFU/mL. After rinsing, cells on these surfaces were enumerated by conventional plating as well as direct counting via scanning electron microscopy (SEM). Compared with the negative control, the positive control and silica aerogel led to a reduced number of salmonellae by 1.0 ± 0.1 log units (87.41 % - 92.05 %) and by 3.1 ± 0.3 log units (99.84 % - 99.96 %), respectively (p < 0.05). The log reductions in the number of Listeria innocua were 1.1 ± 0.1 (90.00 % - 93.69 %) and 3.2 ± 0.3 (99.87 % - 99.97 %) for the positive control and silica aerogel, respectively (p < 0.05). Additional bacterial proliferation studies revealed that bacterial anti-adhesion properties not antibacterial effects were responsible for the observed reductions. Overall, bacterial anti-adhesion property as well as other distinctive properties such as superior thermal insulation and ultra-lightweight make hydrophobically-modified silica aerogel an attractive candidate as a novel food-contact surface.

Coatings with graded mechanical and compositional properties have recently attracted the attention of researchers in the development of biomaterials and tissue engineering supports. Two- and three-dimensional materials featuring discontinuous properties were shown to trigger a spatially-defined cell behaivior or, alternatively, enhance the integration of synthetic matrices with complex biological systems. In natural “graded” systems, in fact, properties of materials such as swelling, stiffness, density and composition are varied in order to respond to external stimuli, either mechanical, physical or chemical. Examples are provided in joint systems where cartilage presents graded composition, water content and mechanical properties in order to both lubricate and support mechanical stress.
We aimed to fabricate graded, quasi-3D polymeric architectures by sequential surface-initiated polymerizations (SIPs). This controlled fabrication relies on an exquisite polymer chemistry-cantered method featuring the grafting of layered polymeric films from immobilized initiators on flat surfaces. Sequential termination/re-initiation steps in selective solvent environments proved as an efficient strategy to graft multi-layer structures with well-defined chemical and physical properties. The use of selective solvent media for the sequential grafting process assured an efficient confining of SIP from the interface of the first layer and avoided re-initiation from the underlying unreacted substrate-bounded initiators. Thus the “bottom” layer acts as a resist for the propagating “top” one, and enables the synthesis of graded polymer films with well- defined vertical compositions.
Following this approach we report the synthesis and characterization of responsive layered films presenting alternated chemistries and different brush architectures. Namely covalently crosslinked poly(hydroxyethyl methacrylate) (PHEMA) brush-gel were layered within linear poly-N-(isopropyl acrylamide) (PNIPAM) brushes forming bi- and tri-layer films by sequential surface-initiated atom transfer radical polymerization (SI-ATRP). Selective solvent environments during the sequential grafting of PNIPAM and PHEMA were allowed by the thermoresponsive behaviour of the former polymer, which, in HEMA-water mixtures, showed a lower critical solution temperature (LCST) below the reaction conditions of 25°C. The mechanical and tribological properties of multi-layered PNIPAM/PHEMA brush/hydrogel films, studied by AFM, showed a strong dependence on layers architecture and responded to the medium temperature. We thus demonstrate how interconnected, layered brush films modulate their properties in a concerted fashion i.e. each layer contributes to the properties of the whole film. We believe that these fabrications can be applied to develop novel polymeric interfaces with fully tunable physical and chemical properties which can mimic the variations typical of natural tissue environments.

Effective anti-fouling coating has attracted much attention due to the increasing demands in various fields, such as biomedical fluid handling, fuel transport, chemic al shielding, marine industry, and so on. Inspirations from nature have paved the way for many revolutionary developments with fascinating functions. In this study, silicon nanopillars with high aspect ratio were fabricated by metal-assisted chemical etching mimicking the 3-D structure of waxy zone in carnivorous pitcher plant. By tuning the density and aspect ratio of nanopillars, the wettability of silicon wafer can be successfully changed from hydrophilic to hydrophobic (CA). In addition, a series of triethoxysilane-based, organic/inorganic hybrid materials were synthesized by sol-gel method for surface modification of silicon nanopillars. The modified Si nano-pillar substrate can achieve superhydrophobicity (CA), extremely low contact angle hysteresis (CAH) (le; 5 degrees) and roll-off angles (le; 2 degrees). Various solvents with low surface tension (le; 27 mJ/m²), such as octane, hexadecane, acetic acid, vegetable oil, were evaluated and results showed oleophobic property (CAsim;120 degrees) due to the surface modification via spin coating and thermal vapor deposition. The dynamic behavior of liquid droplets on the liquid/solid interface was investigated and recorded by high speed camera (Fastec Imaging Co., TS4). The combination of nanostructures and efficient omniphobic coatings by simple processes can significantly improve the performance of liquid repellency comparing to the conventional approaches and shed light on the novel development of anti-fouling and multifunctional surfaces. This research is funded by Material and Chemical Research Laboratories of Industrial Technology Research Institute and the Ministry of Science and Technology, Taiwan (MOST 101-2628-E-007-017-MY3).

DNA is one of the most fascinating biomaterials today and it is a promising molecule for applications in molecular electronics. Moreover, DNA is currently used in the diagnosis of many diseases and a clear picture of the conductance of these molecules could open the doors for the design of diagnostic tools that could be read electronically, thus improving the sensitivity and reducing costs. Although results of DNA conductance reported in the literature span a huge range and differ by orders of magnitude, some consensus has been achieved in the charge transport mechanisms, where tunneling is found in short molecules and hopping in longer DNA molecules. However, to date, conductance modulation by controlling the structure in different DNA forms has not been systematically studied. The B-form typical for dsDNA is a right-handed double helix and it has been extensively studied. The A-form is the prototypical structure for dsRNA and it can be induced in dsDNA by dehydration.
Herein we report conductance measurements of amino-functionalized short dsDNA molecules using the STM-break junction method. Briefly, a gold STM tip is brought into contact with a gold electrode and then retracted with the molecule present in solution while the current between the electrodes is recorded. This process is repeated thousands of times and results can be used to obtain conductance histograms. We study dsDNA conductance as function of length and structure. The structure is changed from B-form to A-form by adding ethanol during the experiment. Results demonstrate that A-form dsDNA is ~10 times more conductive than B-form in these GC rich sequences. This large conductance increase is fully reversible, and by controlling the chemical environment, the conductance can be repeatedly switched between the two values. Length dependent conductance studies of the two conformations suggest that hopping is the dominant charge transport mechanism in these guanine-rich sequences. Ab initio electronic structure calculations coupled with Green&’s Function transport calculations of the two conformations indicate that the HOMO is extended through the entire chain in the A-form DNA case, and this extended orbital results in a higher conductance.
We also report results from electrochemical experiments with methylene blue functionalized DNA in order to obtain information about the interfacial charge transfer resistance and kinetics for the different structures in oligonucleotides ensembles covalently bound to electrodes. These results correlate the structural modulation of single molecule conductance with the electrochemical signal of DNA functionalized electrodes and demonstrate DNA as a promising molecular switch for molecular electronics applications. Additionally, these results could pave the way for the design of sensors that benefit from this conductance-structure modulation in the differential detection of oligonucleotides with identical sequences but different structures.

A variety of sensing materials with specific molecular recognitions are widely investigated to develop high performance bio- or chemical sensors/detection devices. In this study, we introduce a sensing molecule, “Molecularly imprinted polymer (MIP),” which can be used to fabricate advanced bio-sensors. MIP&’s system contains molecular recognition sites, “molecular binding sites,” which are created by “a molecular imprinted technique” to create molecular recognition functions in cross-linked network. However, sensitivity of MIP&’s based bio-sensors limits for practical applications due to the low sensitivity. To achieve a high sensitivity in MIP&’s based sensors, synthesis of “high affinity receptor sites,” such as “monoclonal particles” is a key objective. In previous studies, affinity distribution plots showed that “high affinity binding sites” were obtained when the number of binding sites per particle decreased. This means that smaller particles are expected to have higher affinity binding sites compared to larger particles. The results motivated us to produce small-sized MIP&’s particles for achieving high sensitivity. We employed a novel synthetic technique, a microfluidic synthesis of MIP&’s particles. However, the microfluidic synthesis gave us a difficulty, especially a collection of MIP&’s particles from PDMS-based outlet microchannels due to a sticking problem. Thus, we employed a photocured approach, which fabricated small patterns of photocured MIP on a glass slide. We used a photomask with micro-sized patterns to easily fabricate and then collect MIP&’s particles. The resulting MIP&’s pattern was observed by SEM. Micro-sized MIP&’s particles were collected from the glass surface by scratching off the photocured MIP&’s patterns.

We fabricate free-standing nickel inverse opal membranes in various pore sizes and large areas. With an orderly honeycomb structure and interconnected pore channels, the free-standing inverse opal membranes, after hydrophilic treatments, are experimented for filtering modified HIV virus. The preliminary results indicate successful separation of HIV virus from the median when the pore channel size is below 50 nm. To validate the separation effect, a particular group of cells, which emits fluorescent light once infected, is introduced to the filtered median, followed by the development of the marker cells in a dark room. We determine that the filtered median is clear of any residual virus. In order to reuse the nickel inverse opal membrane, we have explored several innovative ways to eradicate the entrapped virus.

The miniaturization of devices down to the nanoscale plays a major role in semiconductor manufacturing for biosensor applications. Increasing the interactions that occur at the nanoscale offers a number of advantages including signal amplification, improved sensitivity, selectivity, speed and a broad sensing scheme focusing on a specific target.
In this work, micro- and nano-scale transistor devices based on polymeric semiconductor materials (PEDOT:PSS) as an active channel are presented, which are used to study the cell-substrate adhesion by impedance spectroscopy. In order to improve their performance a fabrication process for nano-sized interdigitated transducer electrodes (IDEs) was developed and first devices were tested compared to their micro-scale counterparts. Here we present the fabrication of a nanoimprint lithography (NIL) mold and wafer scale fabrication of an IDE array with an established protocol based on NIL. A NIL mold with IDE structures with finger width and space from 200-300 nm was designed. Definition of the structures was performed on a 4” Si/SiO₂ (200 nm) wafer by electron beam lithography. Structure transfer was performed by dry etching of SiO₂ with a CHF3 plasma. For the fabrication of metal nanostructures a bi-layer resist lift-off technique (mr-I7020 R on LOR3A) was used. NIL was applied for the definition of the structures followed by plasma etching of mr-I7020 R and wet isotropic etching of LOR 3A to create an undercut for the subsequent lift-off after e-beam evaporation of a metal layer (Au on a Ti adhesion layer).
An optical mask was designed for the integration of contact lines in order to determine the electrical characteristics of the IDE array and first devices were successfully used for the analysis of cell-substrate adhesion.

Widespread interest in producing bio-inspired materials underpins the renewed attention of natural silk materials because they exhibit extraordinary mechanical strength, toughness and offer promise in medicine for vaccine and drug delivery. For example, many vaccines and drugs require constant refrigeration prior to use because they must retain the proper folded shape to function properly. Silk proteins can be employed to stabilize the drugs by locking in the structure (see e.g. Stabilization of vaccines and antibiotics in silk and eliminating the cold chain, Kaplan, PNAS 109, 11981 (2012)). The mechanical properties of many similar biomaterials are a direct reflection of an abundance of non-covalent (i.e. weak) interacting ions, which play a critical role in the assembly and performance of bio-structures like F-actin in muscles, tubulin in the cytoskeleton of cells, viral capsids, and silk. Two main factors that are critical for understanding silks: the nanoscale semicrystalline folding structure, which adds strength and toughness, as well as the degree of hydration of the disordered fraction, which acts to modify these properties. Understanding and controlling these two principal factors are the key to the functionality of protein elastomers, and render silk an ideal model protein for (bio)material design. We describe our investigation of the preparation, characterization and Inelastic Neutron Scattering (INS) studies of natural Bombyx mori silk fibroin proteins derived from natural silkworm cocoons. We present the results of in situ methods for monitoring/controlling the hydration and solvent levels using microbalance techniques. The bulk of these studies employed proteins deposited using electro spinning techniques. These technical improvements were incorporated into our INS studies and have enabled us to begin to reveal interesting microscopic dynamics heretofore not investigated or reported. We will present the current status of our INS findings including preliminary results of the effects of water and methanol interaction and preliminary modeling of the protein dynamics.

Drug administration through oral route is deemed as one of the most hassle-free, convenient delivery methods due to its benign need for simple instructions and the lack of necessity of medical professionals during the intake process. The major challenges in oral drug delivery system include rapid release of drugs at gastric pH for stomach-specific drugs, destabilizing effects at low pH against stomach and the subsequent rapid release behavior for intestine-specific drugs, and variable dose requirements. They need to be overcome for the enhancement of safety and efficacy of drugs. We propose a one-stop solution in oral drug delivery system, which can target the stomach as well as different locations in gastrointestinal tract (GI) with variable loading capacities by exploiting the specific architecture of particles and utilizing the pH responsiveness property by optimizing the chemical composition of the materials.
Specifically, we developed a new all-in-one oral drug delivery system targeting stomach, ileum and jejunum in intestine, and tested their stabilizing effect and controlled release behavior in vitro using model drugs such as sulforhodamine b dye, fluorescent nanoparticles (100nm), and bovine serum albumin. To demonstrate the concept, first, the relationship between drug release rate and incubation time at different pH was established to estimate a drug release profile and to address the efficiency of the delivery system. Quantitative analysis was performed by measuring fluorescence intensities using a fluorometer. Second, the morphological variation of the delivery systems at different test conditions was monitored using transmission electron microscopy (TEM), scanning electron microscopy, and fluorescence microscopy to correlate with drug release behavior. Subsequently, structural stability of biomolecular drugs was evaluated from the analysis of spectral data obtained from intrinsic fluorescence, 8-anilino-1-naphthalenesulfonic acid (ANS) fluorescence, and circular dichroism (CD).
The preliminary analysis shows that our drug delivery system exhibits high drug loading efficiency and at the same time, enables realization of our proof-of-concept target-specific delivery system in response to pH changes in GI tract. Although our study was limited to in vitro experiments, future research will be focused on in vivo demonstration in mice. Overall, this research is expected to contribute to the developments of highly efficient drug delivery systems for orally less stable drugs.

Influenza virus has been responsible for a major respiratory disease. Recent spread of the extremely pathogenic avian and swine influenza viruses paint a morbid picture of the emergence of a more lethal influenza virus. Currently, hypodermic needles are being dominantly employed in universal vaccination. However, inherent limitations such as the need of highly skilled healthcare workers, high cost, and safety concerns over needle reuse render it undesirable in developing nations. As an alternative, solid oral vaccine has emerged as a promising platform due to potential advantages like generation of both mucosal and systemic humoral immune responses, no bio hazardous waste, no prerequisite for cold supply chain for transportation/storage, convenient stockpiling due to solid formulation, long shelf life, and self-administration. Despite these potential advantages of oral vaccines, they are still elusive to commercialization due to instability in gastric environment.
Our research goal is to develop a novel oral vaccine delivery vehicle, which can sense pH change of the environment. To this end, we successfully fabricated a microparticle-based vaccine delivery system, which can protect and release vaccines in response to different pH of stomach and intestine, respectively. In this presentation, we will report a comprehensive methodology to fabricate microparticles and corresponding effect on the vaccine stability. As a systematic approach, concept testing was performed to identify any potential design problems or challenges associated with microparticles prior to their application to real vaccine. Briefly, microparticles were synthesized in oil-in-water emulsion method using PMAA-based anionic copolymers. Combined effects of temperature and solvent composition/evaporation conditions on microparticle characteristics were investigated, which were then used to find key process parameters to make optimally functioning pH-responsive microparticles. Morphological change of the microparticles was shown to be important to maintain high level of antigenic stability from in vitro experiments using model drugs (bovine serum albumin, 100nm polystyrene nanoparticles, and sulforhodamine b). Size and morphology of microparticles were examined with transmission electron microscopy, scanning electron microscopy, and dynamic light scattering at gastric/intestine pH conditions. Quantitative analysis of the loading efficiency and time dependent release profile of model drugs were performed using a fluorometer and fluorescence microscopy. Finally, these proof-of-concept data were confirmed using inactivated A/PR/8/34 (H1N1) influenza A virus vaccine by measuring functional hemagglutinin activity at different test conditions. Although our efforts were aimed at the demonstration of concept in vitro, this work is expected to contribute to development of universal platform for oral vaccines.

DNA:RNA hybrids are important biological components and are integral to the processes of DNA replication, transcription and reverse transcription. However, little is known about charge transport though this mixed oligomer. From our measurement results, these hybrid oligonucleotide pairs have significantly different structure than the common double strand DNA (dsDNA). As such, the charge transport properties of DNA:RNA hybrids are expected to be substantially different than dsDNA. In this work, the conductance of individual DNA:RNA hybrids is measured, and we systematically study the transport properties of the DNA:RNA by changing both the length and sequence of the hybrid pair. We also compare these results to the equivalent dsDNA duplex sequences to obtain fundamental insight into the conductance properties of these important biological systems.
In this work, the conductance of the oligonucleotide duplexes is directly measured using the Scanning Tunneling Microscope (STM)—break junction technique in aqueous solutions. This approach, which has previously been used to obtain reproducible conductance values for dsDNA has been adopted to directly measure individual DNA:RNA hybrid duplexes by linking them in between the tip and substrate in an STM. With this setup, thousands of individual conductance measurements can be obtained rapidly for statistical analysis, thus allowing the most probable conductance of a single molecule to be determined. In this work, we performed measurements on sequences with various numbers of G:C or A:T base pairs. With G:C rich sequences, we found the conductance of DNA:RNA hybrid is ~10 times higher than the dsDNA duplex with the same sequence. Yet with poly A:T sequences, the conductance of DNA:RNA hybrid is almost length independent. Thus, this result provides us a better understanding of the fundamental charge transport mechanisms in DNA:RNA hybrids. Moreover, beyond these fundamental studies, we will also present measurements of biologically relevant RNA sequences when hybridized to their DNA compliment. The measurement of these hybrids may provide the basis for developing label-free, RNA-based, electrical diagnostic tools that exploit the electrical properties of the oligonucleotide duplexes.

Magnetic Materials has gained a lot interests in cancer therapy with AC magnetic hyperthermia. Among all the magnetic materials, Fe₃O₄ nanoparticles (NPs) with various advantages such as good biocompatibility, long circulation time, etc., become one of the most popular research areas for magnetic hyperthermia application. However, the magnetic hyperthermia performance of Fe₃O₄ NPs also varies with shape, size and magnetic anisotropy. Octahedral Fe₃O₄ NPs are known to possess high magnetic anisotropy and are very good candidate for magnetic hyperthermia application. Here we reported a detailed study of octahedral Fe₃O₄ NPs for magnetic hyperthermia application. We have synthesized the NPs with sizes ranging from 10 nm to 200 nm. The Fe3O4 NPs were transferred into water phase with cetyl trimethylammonium bromide and thereafter studied for magnetic hyperthermia. Our results showed that octahedral Fe3O4 NPs exhibited very high specific absorption rate (SAR) value with a wide range of particle sizes. Hysteresis loss of Fe₃O₄ NPs in gel was investigated using vibrating sample magnetometer (VSM) at different magnetic field to explain the SAR values. Moreover, our simulation results correspond quite well with experimental data.

Growing evidence indicate that the morphology of material surface has significant effects upon stem cell fate, suggesting that physical impacts may have important effects as chemical factors on stem cell differentiation. Thus controlling the surrounding physical microenviroment may regulate stem cells fate for further applications.
Currently, studies on stress-induced spreading and differentiation in embryonic stem cells have shown that threshold cell deformation is the key point for triggering spreading responses (Chowdhury, Na et al. 2010).Besides, the patterns with nanoscale disorder structure could induce mesenchymal stem cells differentiation into osteogenic lineage without the assistance of soluble chemical factors (Dalby, Gadegaard et al. 2007). Another researchers have got a similar result by modeling the structural changes in extracellular matrix using nanohelical structures, demonstrating human mesenchymal stem cell differentiated in response to physical microenviroment (Das, Zouani et al. 2013). At nanoscale, surface features of material are several orders of magnitude below that of the cells, and it may therefore be possible to target receptor-driven pathways to control the status of the cells.
In our study, we focus on how physical factors regulate mesenchymal stem cell fate, including surface charge, stress and nanoscale topography. This will have great potentials in tissue regeneration and cell therapies.
References
Chowdhury, F., S. Na, D. Li, Y. C. Poh, T. S. Tanaka, F. Wang and N. Wang (2010). "Material properties of the cell dictate stress-induced spreading and differentiation in embryonic stem cells." Nat Mater9(1): 82-88.
Dalby, M. J., N. Gadegaard, R. Tare, A. Andar, M. O. Riehle, P. Herzyk, C. D. Wilkinson and R. O. Oreffo (2007). "The control of human mesenchymal cell differentiation using nanoscale symmetry and disorder." Nat Mater6(12): 997-1003.
Das, R. K., O. F. Zouani, C. Labrugere, R. Oda and M. C. Durrieu (2013). "Influence of Nanohelical Shape and Periodicity on Stem Cell Fate." Acs Nano7(4): 3351-3361.

Since 1987, tris (8-hydroxyquinoline) aluminum (Alq₃) was first designed by Tang and Van Slyke, several researches have been reported to apply this kind of green-fluorescent material in bio-imaging and sensing systems. Due to both covalent and non-covalent interactions, several kinds of metal organic frameworks (3-D hexagonal Alq₃ microrod complex integrated with single strand DNA) were fabricated by our group through chemical reactions and re-precipitation process. When this DNA-guided Alq₃ complex was combined with target DNA (perfect match), a significant increasing of photoluminescence was observed. However, under same environment conditions, 1-mer mismatch and random DNAs showed negligible change in photoluminescence. In addition, surface morphology of perfect matched DNA-guided Alq₃ complex was more embossed than other DNA-guided Alq₃ complex due to adjoining of DNA and Alq₃ particles. We also detected specific peaks by XRD analysis of radial dimension of Duplex DNA. As a result, this metal organic structure can be used as a cornerstone for undermining the potential use of OLED materials in biological interaction events.

In our recent experimental research, we have demonstrated the influence of electric field on cell functionality in vitro. In particular, cell morphological changes and cell proliferation are significantly enhanced over a narrow range of electric field stimulation conditions. In an effort to rationalise such experimental observations, we provide here a theoretical model proposing the development of bioelectric stress field induced cell deformation. A single cell is modelled as a double layered membrane separating the cell culture medium and the cytoplasm with different dielectric properties. This system is linearized by invoking Debye-Huckel approximation of the Poisson-Boltzmann equation. With appropriate boundary conditions, the system is solved to obtain intracellular and extracellular Maxwell stress as a function of multiple parameters like cell size, intracellular and extracellular permittivity and electric field strength. Under representative conditions of electric field stimulation (100 V/m, DC), the stresses experienced at the membrane are around 100mPa, a value much higher (5 orders) than the stress at a membrane without charge density. Based on the stresses, we predict shape changes of cell membrane by approximating the deformation amplitude under the influence of electric field. When a cell under the influence of E-field is considered to be at the interface with a conducting substrate, the stress moments predict that the deformation will be prolate (stretching in the direction of E-field), whereas on an insulating substrate, similar analysis based on the stress moments predict that the deformation will result in an oblate shape (stretching perpendicular to the direction of E-field). Even though the deformation behaviour is qualitatively different, the stress and the amount of deformation remain comparable when cell is considered on an insulating substrate or on a conducting substrate. The biophysical significance of the presented theoretical model is explained in terms of cell morphological changes and enhanced cell signalling processes due to the local convective transport in the presence of external electric field.

Protein Immunohistochemistry is widely used in medical science. Integration of the probe and target biomolecules generally results from one or more non-covalent interactions: hydrophobic/hydrophilic interactions, ionic bond, positive/negative charge, aromatic π-π stacking and other van der Waals forces. Therein nonspecific binding with similar interaction performance as specific binding in protein immunohistochemistry can produce high background noises, resulting in inconclusive target elucidation that hinders interpretation. Due to nano-fabrication, several shapes of CdSe with high quantum yield were designed and used by our group as biomolecules substrates, including sphere quantum dots (0D) and tetrapod (3D). Results show the detection on four-legged substrate has higher specificity due to its larger molecular volume. In addition, as the non-covalent interactions can be influenced by environmental conditions, we controlled specially the fluid velocity in vortex field as well as the common conditions, such as block agent (ex. BSA), temperature, pH, salt conditions and binding time. Interestingly, higher Accuracy appeared at the high fluid velocity, which means nonspecific binding can be largely replaced by specific binding due to molecular physical interactions. Furthermore, some other kinds of biomolecules (different protein types or same type from different species and DNA with different sequence) were also tested under their optimal conditions to have an overall conclusion.

The development of new materials for use in energy and environmental applications is of great interest, in particular in the areas of gas separation and carbon capture. The dipeptides are one class of organic molecules that offer an attractive possibility in such areas, because they form open hexagonal crystalline structures (space group P6₁) with quasi one-dimensional channels of tunable pore diameters in the range 3-6 Å. Previous studies indicate that these molecular crystals exhibit selective adsorption, as well as, water and gas transport properties. The selective permeability and transport properties of these biomaterials are believed to result from collective vibrations of the crystal structure that are coupled to the motions of the guest molecules within the channels. Zeolites with 1D channels of comparable diameter have been identified for use in methane/carbon dioxide separation and purification even at low pressure. Current studies are focused on L-Isoleucyl-L-Valine (IV) and L-Valyl-L-Alanine (VA), which have highly hydrophobic walls as a result of the inward pointing methyl groups that line the channels. High-resolution methane adsorption isotherms carried out for IV and VA at low pressure exhibit remarkable adsorption properties with IV exhibiting an adsorption capacity of 20 cm³ (STP)/g. Recent results on Metal Organic Frameworks (MOFs) suggest the selective adsorption properties might result from a close match of the gas size and the channel diameter of the dipeptides, due to a sizable van der Waals interaction between the hydrophobic walls and the methane adsorbate. In order to more precisely characterize the dipeptide methane interaction, high-resolution Inelastic Neutron Scattering measurements were performed at the Spallation Neutron Source (BASIS spectrometer). These studies showed clear rotational tunneling peaks for the IV-methane system, which can be used to gauge the rotational barrier and details concerning the potential energy surface. Some evidence that the flexibility and dynamical motion within the biomaterials channels play a significant role in the adsorption properties will also be presented.

Transplantation of human neural stem cells (hNSCs) has been proposed as an effective therapeutic strategy for the central nervous system (CNS) repair, restoring damaged tissue and rescuing lost functions of CNS.
hNSCs are known to form spherical clones -neurosphere,- in vitro by cell migration and proliferation. The different cellular states are represented by neurospheres due to the unique internal microenvironments in the three-dimensional (3D) cell cluster. Conventional two-dimensional (2D) culture system can&’t control size of neurospheres. Therefore, it is difficult to study with heterogenous size of neurosphere in 2D culture system. Here, newly designed 3D cell culture system for hNSCs in introduced to control of neurosphere formation.
In the previous study, our group fabricates a quasi-spherical droplet cell culture system -called “Spheroform”- for the formation of 3D cellular spheroid. The construction of 3D environment resulted in an enhancement of therapeutic and biological functions. This cell culture system utilizes dot patterning of several hundred micrometers on the superhydrophobic TiO2 coating surface by poly-dopamine which is self-polymerized mussel-inspired catechol-amine on any surface.
We confirm that the 3D cell culture system can control variety size of neurosphere. We expect that the neurosphere in spheroform can exhibit different biological outputs such as cell viability, gene delivery efficiency and secretion rate of cell signal factors. This 3D cell culture system will possibly contribute to provide platform technique that mimic in vivo hMSC cellular states by controlled neurosphere formation in vitro.

The use of porous solid materials in widely different fields of science can be easily associated with the high level of scientific and technological development currently observed. Significant advances in the ability of manufacturing and manipulating porous solids with the most diverse chemical and physical features resulted in the development of materials with remarkable properties, which have not only boosted the traditional application of porous solids as adsorbents or catalysts, but also have expanded their use to a wide range of applications, e.g., microelectronics, sensing and medical diagnosis. This work describes the utilization of a mesoporous silica matrix organofunctionalized with an imidazolium ionic liquid-based alkoxysilane for the synthesis of supported Pt nanoparticles. The resulting material (SiO₂-Imi-Pt), consisting of highly dispersed metallic nanostructures in the porous framework, has shown special ability to act as an artificial enzyme, displaying peroxidase-like catalytic activity. Such property allows the application of this nanomaterial in (bio)sensing devices, here evaluated towards H₂O₂ detection. The utilization of peroxidase enzyme for monitoring hydrogen peroxide is one the most used strategies in biosensing assays once H₂O₂ is a product of several enzymatic reactions and acts as an efficient probe for the determination of distinct analytes, e.g. glucose, uric acid, cholesterol and lactate in blood samples and also in immunoassays which make use of enzyme-labeled antibodies.The proposed artificial system based on supported Pt nanoparticles is able to catalyze the H₂O₂-mediated oxidation of the peroxidase substrate 3,3,5,5-tetramethylbenzidine (TMB) to produce a blue colored product which can be easily identified and quantified. This attribute was explored for the colorimetric detection of H₂O₂ and also for glucose sensing. In the last case, the glucose oxidase enzyme (GOx) was immobilized on the surface of SiO₂-Imi-Pt. Upon addition of glucose and TMB, a cascade reaction initiates with the oxidation of glucose, formation of H₂O₂ (enzymatic - GOx) and the subsequent H₂O₂-mediated catalytic oxidation of TMB on the surface of the metallic nanostructure. The catalytic properties of the SiO₂-Imi-Pt indicate that this nanomaterial is potentially promising for the development of biosensing devices as a peroxidase mimetic system.

Nanopores are used extensively for studying the single molecule translation and can be used for DNA sequencing, single molecule detection and understanding. Nanopore technique uses electrophoretic forces to drive molecules through a nano scale pore embedded in an insulating membrane. In this method voltage is applied to force charged molecules to pass nanopore and the blockage ionic current change is monitored caused by molecule&’s translocation between two conductive electrolytes through the pore. Nanopore could be formed by pore forming protein (biological pore) or fabricated on insulating membrane (solid- state pore) or hybrid of both. Despite of pore class, its characteristic will affect sensory behavior of nano scale aperture. Pore&’s surface charge and conductance could modulate the pore/ molecule interaction and control the translocation time and capture rate.
In this work, we will analyze nano pore&’s ionic conductance in relation to its size. 25 nm low stress LPCVD silicon nitride membrane (2mu;m×2mu;m) formed by backside anisotropic etching of silicon. Nanopores of 5-50nm were drilled on membrane using Hitachi NB-5000 FIB/SEM. Pore size have been optimized by changing the beam&’s dwell, frame time and aperture size. Piranha cleaned chips clamped between two custom designed Plexiglas holders. Electrical measurements are carried out by connecting Ag/AgCl electrodes to KCl buffer chamber, which is connected to electrolyte chamber via Agar salt bridge. Axopatch 200B patch-clamp amplifier and planner lipid bilayer workstation (warner instruments) coupled with Axon Digidata 1440 digitizer and Bessel filter have been utilized to record ionic current. Whole setup was insulated in Faraday cage and vibration isolation table.
Conductance measurements were performed in 1M KCl-10mM Tris at pH 8.0 as both Cis and Trans electrolyte. Voltage of -0.1 to 0.1V is applied and ionic current across the pore with different sizes is recorded. Pore conductance versus pore diameter will be reported and will be modeled based on its shape. To study the conductance change in presence of single molecule and its effect on translocation, conductance and dwell time will be reported upon insertion of single strand DNA of 749 and 275 base lengths.

In order for gene therapy to be used in clinical settings, safe and efficacious delivery systems must be developed that simultaneously address numerous extra- and intracellular challenges. Non-viral delivery systems, which typically employ cationic lipids or polymers to complex and condense DNA through attractive electrostatic forces, are easier to construct and have superior safety profiles when compared to viral vectors. Despite numerous efforts to resolve their limitations, however, most lipo- and polyplexes suffer from low DNA packaging efficiency, modest targeting specificity, and a high degree of undesired cytotoxicity. To this end, we have developed a novel nanoparticle-based transfection reagent: mesoporous silica nanoparticles (MSNPs) wrapped in supported lipid bilayers (SLBs). We have found that the bio/nano interface between the MSNP and SLB results in unique biophysical properties that are not possessed by either MSNPs or liposomes alone; one such property is the high degree of SLB fluidity and stability, which allows us to achieve high binding affinities when extremely low densities (<10 molecules/nanoparticle) of targeting ligands are conjugated to the SLB. These high binding affinities allow us to selectively deliver a plasmid (pZG1) that encodes the reporter protein, ZsGreen, to hepatocellular carcinoma (HCC) cells with nearly 100% efficiency. Specifically, we use histones to package pZG1 into highly compact nanoparticles that are subsequently modified with a nuclear localization sequence (NLS) and incorporated within the MSNP pores. Zwitterionic liposomes are then fused to pZG1-loaded cores to form a SLB, which we modify with a targeting peptide (MC40) that binds to many types of HCC and with an endosomolytic peptide that promotes endosomal escape. We have shown that our MSNP/SLB nanoparticles have a 100-fold higher capacity for DNA than similarly-sized lipoplexes and retain histone-packaged pZG1 for > 4 weeks when exposed to blood at 37°C. Furthermore, MC40-targeted MSNP/SLBs have a 10³-fold higher affinity for HCC (Hep3B) than for untransformed hepatocytes and are selectively internalized by Hep3B via receptor-mediated endocytosis. Endosome acidification destabilizes the SLB and protonates the endosomolytic peptide, both of which promote efficient cytosolic dispersion of histone-packaged pZG1, while the NLS induces nuclear accumulation and enables transfection of nearly 100% of dividing and non-dividing Hep3B cells, which express ZsGreen for up to 4 weeks. We are currently assessing the ability of pZG1-loaded MSNP/SLBs to selectively transfect human cancer cells in a murine xenograft model; preliminary findings suggest that target cells express 50-fold more ZsGreen when mice are injected with MC40-targeted MSNP/SLBs versus MC40-targeted DOTAP/DOPE lipoplexes. In conclusion, we believe MSNP/SLBs possess the capacity, specificity, and stability necessary to overcome many of the barriers to effective gene therapy.

The aim of this research was to develop a sensor platform that enables the detection of fungicide residues in a facile manner. Through the use of electrochemical impedance spectroscopy (EIS), this work shows a simple process for the fabrication of a sensor platform which is able to be modified for the detection of multiple target analytes. By employing EIS we were able to demonstrate a sensor which could rapidly determine the concentration of the target fungicide present in a grape extract which simulated real-world conditions. In order to test the specificity and sensitivity of the sensor and its ability to work for the proposed application, the samples of grape extract were spiked with the fungicides azoxystrobin, trifloxystrobin, and pyraclostrobin.
Fungicides are a type of pesticide used for the control and prevention of fungal growth in many commercial crops. Current methods used for the detection of fungicide residues in commercial crop samples include both high performance liquid chromatography and gas chromatography mass spectrometry. These methods are time consuming and require that the crop sample be sent to a laboratory for processing and testing by trained laboratory personal. This makes these methods both costly and time consuming. A sensor which is able to rapidly determine the concentration of fungicides in a crop sample at the point of test would reduce cost and expedite the process of moving the crop from the farm to market.
To create a sensor which was able to perform the rapid point of test analysis that was needed, we functionalized gold electrodes which were patterned on a printed circuit board (PCB). The gold electrodes were first coated with DSP to form a linker molecule layer. This linker utilized thiol binding to adhere to the gold and contained an NHS ester group on the free end which is capable of binding free amine groups of the antibodies used to capture the target fungicides. This simple approach provided the ability to easily switch between antibodies for targeting multiple fungicides.
This work demonstrated sensors which were able to detect the presence of fungicides down to the pg/mL range, which was well below the needed level of detection. We were able to achieve these results in less than 15 minutes, showing that this robust sensor platform could work for the rapid, point of test detection of fungicides.

The properties of molybdenite (MoS₂) make it useful in many applications. Besides its main consumption as a dry lubricant and catalyst for desulfurization, recent research implies that it has potential in the fabrication of ultrasensitive and switchable transistors, light emitting diodes and solar cells. The enlarged application areas result in higher demand for molybdenum worldwide. Even though there are principle mines producing molybdenite, a remarkable amount is found as a trace element in copper and tungsten mines. However, the separation and purification of this valuable and trace element requires extensive processing and mostly left over in tailings. A targeted purification approach could be applied in order to recover such valuable elements from acid mine drainages, tailing ponds and polluted water sources in a more effective and environmentally friendly way.
As a general strategy, we aim to identify biomolecules (peptides) with specific recognition ability to individual metals and coat magnetic particles with these peptides for the production of smart biomaterials. These biomaterials would specifically bind to the targeted metal and would be separated/recovered by applying a magnetic field. Additionally, the particles would be recycled after the dissociation of the metal from smart biomaterial.
Here, we selected MoS₂ binding peptides from a 12-mer peptide library using phage display techniques. Among the 43 peptides selected, 11 peptides exhibiting more than 80% binding affinity have been classified as strong binders. The binding strength of some of the strong binding peptides was tested in various pHs between pH3.0 and pH9.0. Almost all of the peptides retained their binding ability in that wide range of pH, which is promising for their potential utilization in various tailing ponds. Additionally, the peptides were evaluated for cross-specifity with other materials (Al₂O₃, Fe₂O₃, CaO, MgO, SiO₂, Zn, Cu, S, Tungsten, Graphite). None of the strong binders showed a specific affinity of greater than 10% for these materials except Graphite. Meanwhile, a variety of Fe₃O₄ magnetic nanoparticles were synthesized and surface functionalization with peptides was initiated. The next step will be monitoring smart biomaterials interaction with MoS₂ particles and their separation under magnetic field.

We incorporated natural extracts having antibacterial property into nanoclays through nano-bio hybridization method. Four kinds of mineral materials such as layered double hydroxide (LDH) illite (IL), holrait (HO) and scoria (SC) were selected as nanoclay candidates. In order to provide porosity to nanoclays, LDH was thermally treated at 400^oC, and the other nanoclays were treated with 6M HCl. Through hydrothermal extraction, we obtained natural extracts of Paeonia suffruicosa Andrews (PS), Agrimonia pilosa Ledeb (AP) and Paeonia japonica var. pilosa Nakai (PJ) which were determined to have antibacterial activity on both Gram positive Bacillus subtilis and Gram negative Escherichia coli bacteria. Four kinds of nanoclays and three natural extracts were hybridized in aqueous media to produce 12 nano-bio hybrids. We evaluated the natural extract contents in each nano-bio hybrids by weighing method and chose hybrids with natural extract above 10 wt% for further study. The power X-ray diffraction patterns, scanning electron microscopic images and zeta potential measurement indicated that natural extracts were mainly adsorbed on the surface of nanoclays. Thermogravimetric and differential thermal analyses exhibited that the combustion of natural extract in the hybrids occurred at lower temperature compared with natural extract alone, suggesting the molecular rearrangement of natural extract in the hybrids. The antibacterial activities of the hybrids were evaluated by bacterial colony count method, showing similar or enhanced antibacterial activity for the hybrids compared with natural extract alone.

Biofilm formation by bacteria on implants may have severe effects on the lifetime of implant devices, also represents a major cause of implant failures and even patient morbidity. . Treating the implant infection by antibiotics is not effective due to their poor penetration rates in biofilms and also the effect bacterial resistance towards multiple antibiotics poses serious concerns. Polymer materials used as delivery systems for antibiotics or metallic ions have also been investigated. The difficulty in engineering these systems is that the release kinetics is complex and that the polymer may mitigate some of the antibacterial properties of the active ingredient, they may also cause to trigger bacterial resistance as well as cytotoxicity issues remains questionable. An optimal design of non-adhesive and infection resistant surface coatings using alternative antimicrobial agents may help to solve the challenges. By designing tunable interfaces, we propose a self-organizing solution that reduces the size of the delivery system to close its theoretical limit. The antimicrobial peptides and proteins are evolutionary conserved constituents of the immune defense of many organisms including insects, plants, and animals. Antimicrobial peptides (AMPs) are known to have high lethality and broad spectrum activity compared to antibiotics. To attach the AMP to the implant, we incorporate them with genetically-engineered peptide for inorganics (GEPI) tags that uses non-covalent interactions specific to the material of the implant surface. To preserve the function and orientation of each attached peptide unit, we specifically engineer spacer sequences to enhance the chimeric functionality. The engineered spacers are also peptide sequences that may not interfere with the displayed functions yet may allow to display multiple functions. These peptides are considered as part of engineering the interfaces by expanding the de nova design approaches for creating the chimeric peptides that may have wide spectrum application ranges. The spacer sequence introduces a new level of peptide assembly organization. In our presentation, we will provide several spacers as part of our engineering design. They were designed to be rigid, hydrophilic or to be disordered with selected secondary structures. The efficiency of engineered chimeric peptides both in solution and on implant surfaces, titanium and titanium alloys and zirconia were evaluated in vitro against common oral and orthopedic infectious organisms, S. mutans, S. epidermidis, and E. coli. We greatly acknowledge the funding from NIHR-NIAMS, KU-NFRG, and KU Internal Funds.

TiO₂ and SiO₂ nanomaterials are the most widely used nanomaterials in food applications as whitening and anticaking agents. Due to the world-wide increasing concerns on potential toxicity of nanomaterials, it is required to evaluate the interaction between food grade nanomaterials and biological substances like electrolyte, sugar molecules and proteins. In order to assess to biological behaviors, we treated food grade TiO₂ and SiO₂ nanomaterials in various solutions such as deionized water, phosphate buffered saline (PBS) and protein or sugar solution (albumin or D-(+)-glucose). The physicochemical properties of nanomaterials in the suspension were evaluated in terms of hydrodynamic size, zeta potential and surface chemistry. Both TiO₂ and SiO₂ nanomaterials showed significant agglomeration in deionized water and PBS, possibly due to the inter-particle interaction and surface adsorption of electrolytes. Albumin was determined to accelerate agglomeration of SiO₂ nanomaterials, but to increase colloidal stability of TiO₂ nanomaterials. Glucose showed reverse results; agglomeration of TiO₂ increase and colloidal stability of SiO₂ increased. From surface chemistry analyses such as FT-IR and XPS, we found that albumin and glucose were not strongly bound to nanomaterials&’ surface showing the preservation of chemical environments around Ti, Si and organic moieties. It could be concluded that the bio-substances can either increase or decrease nanomaterials&’ colloidal stability by simply adsorbing on the surface of nanomaterials.

An urgent need exist for developing handheld devices for rapid, sensitive, and specific detection method for pathogens.[1] Here we demonstrate a rapid detection method for Gram-positive and Gram-negative bacteria using an impedance sensor array functionalized with antimicrobial peptides (AMPs). This impedance sensor screens pathogens in real-time and has comparable sensitivity with current detection methods like polymerase chain reaction (PCR) and immunoassay.[2,3] Functionalized electrodes in array selectively bind to the corresponding bacteria strains, especially the pathogenic strains resulting in variations in the impedance modulus. Impedance variation is used to detect incubated bacterial cell concentration with a resolution of 1 cell/µL. The dynamic range of detection for both Gram-positive and Gram-negative bacteria is found to be 10³-10⁶ cfu mL^-1. Micropatterned electrodes modified with AMPs in an impedimetric array offers an excellent platform for rapid and selective detection of pathogens in contaminated water and food products.
Reference
[1] W.H. Organization, Microbiological Hazards in Fresh Leafy Vegetables and Herbs: Meeting Report, World Health Organization, 2008.
[2] J.D. Slinker, N.B. Muren, A.A. Gorodetsky, J.K. Barton, J. Am. Chem. Soc. 132 (2010) 2769.
[3] P. Arora, A. Sindhu, N. Dilbaghi, A. Chaudhury, Biosens. Bioelectron. 28 (2011) 1.

Natural cotton was dissolved in the room temperature ionic liquid 1-ethyl-3-methyl acetate and wet-jet electrospun to obtain nanoscale cotton fibers having an increased surface area relative to natural cotton fibers. The resulting nano-cotton fibers were esterified with trityl-3-mercaptopropionic acid, which after selective de-tritylation afforded nano-cotton fibers containing reactive thiol functionalities. Silver nanoparticles were next assembled that were covalently attached to these sulfhydryl groups. The microstructure of the resulting nanocomposite was characterized, and the antibacterial activity of the resulting nano-cotton Ag-nanoparticle composite was also studied. This nanocomposite showed significant activity against both Gram-negative and Gram-positive bacteria.

Controlling cell adhesion, migration, proliferation and differentiation is crucial for biomaterials in a number of applications. The adjustment biomimetic surface properties of the material through immobilization of extracellular matrix derived peptide motifs are important to facilitate effective interactions between cell receptors and the surface of artificial scaffolds. These biomimetic surfaces should suppress the protein adsorption from the biological media to which they are exposed to, cancel the non-specific cell-material interactions and, at the same time, elicit a specific cell response to biologically active surface immobilized molecules.
Herein we present a detailed study of dense PEO brushes prepared by end-tethering of hetero-bifunctional PEOs (M_n=2,000-20,000) to polydopamine (PDA)-modified surfaces from a reactive melt. [2,3] The presence of alkyne distal end-group on the PEO chains was used to couple azidopentanoic-bearing peptides utilizing a copper-catalyzed Huisgen azide-alkyne #8222;click#8223; cycloaddition reaction (CuAAC). The peptide surface concentration was tuned from complete saturation of the PEO surface with peptides (1.7×10⁵ fmolmiddot;cm^-2) to values still relevant for cell adhesion studies (<6.0×10² fmolmiddot;cm^-2). Detailed physico-chemical characterization of the layers was performed using different surface sensitive techniques (spectroscopic ellipsometry, contact angle goniometry, electro-kinetic measurements, infrared reflection-absorption and vibrational sum-frequency-generation spectroscopy). Radioactivity assay and imaging of ¹²⁵I-radiolabeled peptide derivatives were used to precisely determine the surface concentration and the macroscopic uniformity of the surface bound peptides. Non-specific protein adsorption was monitored by surface plasmon resonance spectroscopy. The performance of the prepared surfaces was tested in cell cultures of human umbilical vein endothelial cells.
Acknowledgement: Support from the Czech Science Foundation (project: P108/11/1857) and the European Regional Development Fund project #8222;BIOCEV - Biotechnology and Biomedicine Centre of the Academy of Sciences and Charles University“ (CZ.1.05/1.1.00/02.0109) is gratefully acknowledged.
References:
[1] Proks, V. et al., Macromolecular Bioscience 12, 1232 (2012).
[2] Pop-Georgievski, O. et al., Langmuir 28, 14273 (2012).
[3] Pop-Georgievski, O. et. al., Biomacromolecules 12, 3232 (2011).

Textiles have long been known as suitable media which support the growth of microorganisms. These microorganisms negatively affect public health and also degrade the performance of the textile itself. Therefore, antibacterial agents are very important in the textile industry. Antibacterial activity is related to compounds that locally kill bacteria or slow down their growth, without being in general toxic to surrounding tissue. Antibacterial agents are paramount to #64257;ght infectious diseases. Recently studied, boron nano-particles (applied to table grapes) have shown good antimicrobial activity. Boron is effective to control gray mold on table grapes caused by B. cinerea both at room temperature and at 0 °C. The efficacy is positively correlated with the boron concentration in the solution [1].
The antibacterial properties of boron-containing compounds are well known although there are limited studies available on the pure boron nanoparticles. In this study boron nano-particles were characterized in terms of their particle size, shape, stability and surface charge before and after they are applied to textile surfaces to study their impact on bacterial activity in addition to cytotoxicity. It was observed that the boron nano-particles are affective in limiting bacteria growth on both gram-negative and gram-positive species without requiring any stimulation to initiate the antibacterial action. It was also found that the application of boron nano-particles on the textile surfaces through mixing them in hydrophobic finishing solutions helped improve the wettability performance of the textiles while showing no change in the physical and colour fastness properties at an optimal concentration of 0.02 g/100 ml of finishing solution.
References:
1. Qin, G.; Zong, Y.; Chen, Q.; Hua, D.; Tian, S. Inhibitory effect of boron against Botrytis cineria on table grapes and its possible mechanisms of action, International Journal of Food Microbiology, 138, 145-150, 2010.

Polymer coatings are used as adhesives, tissue engineering scaffolds, wound healing materials, and drug delivery systems. Design of rapid fabrication techniques provides avenues for the development of new products with minimal manufacturing and production issues. Microgels are discrete hydrogel microparticles that have previously been used in biomaterial applications due to their tunable mechanical and chemical properties. In this work, we employ microgels composed of the monomer N-isopropylacrylamide, co-monomer acrylic acid, and cross-linker N,N&’-methylenebisacrylamide as well as the polycation, polyethyleneimine to develop biomaterial systems.
Traditionally, polyelectrolyte microgel films have been fabricated via the time-consuming layer-by-layer (LbL) method in which anionic microgels are centrifugally deposited followed by passive adsorption of a polycation for film build-up. We have developed a rapid film fabrication technique wherein the polycation and microgels are mixed together and immediately deposited via centrifugation. This fast method has enabled the development of films with architectures that were not feasible given previous LbL methods. Here, we have fabricated thin microgel films, ultra-thick microgel films, patterned microgel films, gradient films, films composed of additional polymeric building blocks, and even bulk gels.
Materials were characterized via brightfield microscopy, confocal microscopy, scanning electron microscopy (SEM), atomic force microscopy (AFM), AFM nanoindentation, and cone and plate rheometry. AFM images indicate microgel film morphology is consistent with traditional LbL films. Analysis of SEM images indicates exponential film growth occurs. AFM nanoindentation studies reveal that the micro-mechancial properties of the films vary inversely with film thickness. Thick films (~ 50 um) have a young&’s modulus of ~ 1 kPa, two orders of magnitude lower than typical films containing 4% BIS microgels. Thin films (~ 500 nm) have a young&’s modulus ~200 kPa, which is comparable to that of a traditional 4-layer LbL film. Brightfield images of patterned microgel films reveals a high pattern fidelity for micron-scale features. We are currently using patterned films to modulate cell adhesion and spreading for tissue engineering applications. Cell attachment studies demonstrate that fibroblasts preferentially spread in between the microgel film pillars. Additionally, bulk gels can be chemically cross-linked to tune the mechanical properties for development of tissue-engineering scaffolds as demonstrated through AFM nanoindentation studies. Oscillatory rheology measurements of these samples in the linear regime indicate that, regardless of crosslinking, mechanical properties of the materials are similar and reminiscent of an elastic system on the region where experiments were performed. Finally, we are exploring pH responsivity of patterned microgel films for use in wound healing and triggered drug release.

Nanoporous gold (np-Au) thin films, produced by a dissolution-based self-assembly process, offer high effective surface area, electrical conductivity, catalytic activity, tunable pore morphology, and compatibility with conventional micropatterning techniques. These properties are critical for a number of technologically important applications including biosensors, fuel cells, and photonics. In particular, np-Au thin films are an excellent choice for nucleic acid-based bioanalytical sensors owing to their three-dimensional surface and ease of functionalization. The sensor performance can be enhanced by leveraging the probe-target interactions within these nanostructured thin films. Tunable pore morphology serves as a versatile model to investigate the interaction of nucleic acids with the nanopores in different size scales. Having a library of structures with varying morphology may help in expanding the useful dynamic range of detection. Different film morphologies were obtained by annealing the nanoporous films at different temperatures. We report a systematic investigation on influence of varying nanoporous morphology on DNA-methylene blue (MB) interactions and their application for development of DNA biosensors. Np-Au thin film electrodes with sub-micron thicknesses were employed for DNA immobilization and square wave voltammetry was used for characterizing probe grafting density and monitoring target hybridization. The interaction mechanisms of the small redox molecule (methylene blue) with single and double-stranded DNA immobilized onto np-Au surface was electrochemically investigated. Electrochemical current decay and regeneration were influenced by the mode of attachment of MB to the DNA: covalently attached MB displayed no decay in current over repeated cycles of square voltammetry, while the decay in physically attached MB depended on the accessibility of the effective surface. Also, the optimum frequency for extracting the maximum current signal depended on the morphology. The optimal frequency was found to be lower when the surface was less accessible. These two findings provide novel insights into the transport mechanisms of small molecules in np-Au films. The np-Au electrode exhibited a significant enhancement in probe grafting density and about 20-fold higher current for target DNA as compared to a flat gold electrode. In addition, the electrode showed sensitive detection with a dynamic range of 10 nM to 100 nM. An order of magnitude shift in dynamic range of detection was observed with annealed nanoporous films due to increased accessibility into the porous network and hence enhanced hybridization efficiency. We believe this study will pave the way to design optimal sensing electrodes at different size scales for high performance electrochemical biosensors and will have potential applications in point-of-care platforms.

Soft materials such as polymers and fibers are key industries in modern society and play an important role in our daily lives. These materials contribute to our society in a wide range of applications such medical, electronic, and optical devices. It is also expected that in the future soft materials can expand its role in research related to the environment, energy science and green technology such as the separation of oil-water mixtures. Oil-water separation is a worldwide challenge because of the frequent oil spill accidents and increasing industrial oily wasted water. Materials that can effectively separate oil and water are in great demand from the aspects of environmental and economic. Here inspired by the lower surface of the lotus leaves and fish scale we developed a special kind of soft material PEG cryogel membrane with aligned porous structure by the unidirectional technique. This material has special wettability of superhydrophilicity and underwater superoleophobicity which enables them the ability to effectively separate oil-water mixtures including the emulsions with micro-sized oil droplets. This material can separate free oil-water mixtures, the mixture of different oils and water and the emulsified oil-water mixtures with great efficiency of up to 99%. Our separation methodology is solely gravity driven and consequently expected to be high energy-efficient, the underwater superoleophobic interface with low affinity for oil droplets can prevent the material from being contaminated by oils, which makes the recycling of oil and materials easy. This porous soft material with water removing ability has the advantage of energy saving and environmental friendly, because of its special wettability that is opposite to traditional oleophilic materials that it overcomes the easy-fouling and hard-recycling limitations in essence. It is a new attempt to use special soft material in the separation of oil-water mixtures which suggests attractive potential applications in industrial oily waste water treatments and oil spill cleanup.

Nanopore technology has attracted tremendous interest in recent years due to their prominent potential to solve the ever-increasing demands of genome sequencing, protein sequencing and small molecule sensing for disease diagnostic. Two types of nanopores, namely biological nanopore and solid-state nanopore, have been developed and used in DNA sequencing. Each of them has their pros and cons. Specifically, biological nanopores that are made by porin or channel proteins embedded in lipid bilayer have superior selectivity but are generally unstable for long-term measurement. Instead, solid-state nanopores are more durable but lack of specificity. Though a hybrid nanopore that is made by insertion of channel protein on a solid-state nanopore was reported to improve the selectivity of solid-state nanopore, it didn&’t solve the problem of poor long-term stability caused by the protein. Generally, the pore and channel proteins are integral membrane proteins (IMPs) that have hydrophobic surface which is embedded in lipid membrane in their native state. As a result, free molecules of IMPs are unstable in aqueous solution. Thus far, there is no report on a hybrid system that exhibits both selectivity and durability. The challenge lies in maintaining the long-term stability of the pore protein in the working condition, i.e. aqueous environment. Here we propose to use an amphiphilic beta-sheet forming peptide to stabilize the protein. Replacing the role of detergent, the peptide binds to the hydrophobic surface of IMPs and forms a beta-barrel wrapping around the protein. We have purified a variety of IMPs and demonstrated that the peptide-stabilized IMPs remain their structure in detergent-free buffer over a period of 3 months. We have generated solid-state nanopores that have size of tens of nanometer based on silicon nitride membrane. We are currently testing polymer growing on the nanopore to modify the solid-state nanopore properties. The polymers also serve as a cross linker to covalently immobilize the channel protein on the solid-state nanopore. The resulting hybrid system therefore combines the advantage of the selectivity of a biological nanopore and durability of solid-state nanopore.

The surface of a nanomaterial plays one of the most important roles in determining its interaction with a biological system, including the resistance of protein adsorption, blood circulation, tumor targeting, deposition of extracellar matrix, cell attachment, and tissue regeneration. Unfortunately, we often have very little quantitative information about the topmost layer on the surface of a nanomaterial, especially when the nanomaterial is suspended in a liquid medium. In this talk, I will discuss a number methods we recently developed for achieving a quantitaive understanding of the molecular or ionic species adsorbed on the surface of nanoparticles. The techniques include inductively coupled plasma mass spectrometry (ICP-MS), surface-enhanced Raman spectroscopy (SERS), and UV-vis/fluorescence spectroscopy. We have applied these techniques to a number of systems, in an effort to quantify the coverage density of the molecular/ionic species on the surface of a nanoparticle. I will also show a few examples, in which such quantitative information has allowed us to greatly improve the performance of various nanomaterial in biomemdical applications.

Neural electrodes constitute an important tool for monitoring and modulating the electrophysiological activity of the nervous system. A major obstacle in long-term reliability of neural recording electrodes has been the undesired aggregation of glial cells on the surface of the electrode and subsequent death and/or detachment of neurons from the electrode surface. Multifunctional electrode coatings have shown potential for improving electrode-tissue interaction. Nanoporous gold (np-Au), produced by an alloy corrosion process, is a promising candidate for multifunctional neural electrode coatings, due to its in situ drug delivery capability, performance in high-fidelity electrophysiological recordings, and compatibility with microfabrication. This in vitro study on np-Au&’s interaction with cortical astrocyte-neuron cultures reveals that np-Au&’s nanostructure significantly reduces astrocyte coverage (p < 0.001) while maintaining high neuronal coverage (p > 0.9) in comparison to planar gold surfaces. This reduction in astrocytic coverage may be partially responsible for previously-reported high recording fidelity from organotypic slices on np-Au multiple electrode arrays. In this work, we present a systematic study, where chemical and morphological cues are decoupled by a combination of thin film sputtering and atomic layer deposition. First, np-Au films with varying atomic percentages of residual silver (12%, 5%, 3.5%, 2.5%) are tested for potential cytotoxicity. It is seen that residual silver elicits an adverse response from astrocytes above 5% silver. However, since the films used in this study are between 3-5% silver, the reduction seen is not primarily due to leached silver. Based on our data indicating that the increased neuron/astrocyte coverage ratio of np-Au is not due to toxic effects of leached silver on astrocytes, we investigated whether insoluble silver that may be present on the np-Au surface or morphological cues (i.e., np-Au nanostructure) alone dictate differential cortical-cell coverage. In order to decouple these effects, both np-Au and planar gold surfaces were coated with a 2.5 nm-thick conformal coating of aluminum oxide using atomic layer deposition (ALD). The conformal coating created by ALD uniformly masked the metal surface with alumina, thereby altering the surface chemistry of the np-Au without affecting the morphology. The neuron-glia co-cultures grown on the ALD np-Au and np-Au surfaces showed similar neuron coverage (p > 0.6) on both surfaces while astrocyte coverage exhibited similar reductions on both the ALD np-Au and np-Au surfaces (p > 0.03). The results demonstrate that the np-Au surface morphology rather than the chemistry dictates cell type-specific surface coverage. This work identifies np-Au as a novel material for multifunctional neural electrode coatings, as well as a surface that facilitates neuronal enrichment in the absence of cell culture medium supplements that reduce glial overgrowth of cultures.

Biomimetic materials design holds enormous potential for the generation of technologically advanced materials by exploiting the inherently specific and programmable nature of biomolecular interactions. For example, peptides that have specific affinities for materials surfaces have been utilized to control the assembly and interfacial properties of a wide range of abiotic chemistries. The complex structure and functionality native to biomolecules enables one to envision a future in which materials properties can be controlled by designing biotic/abiotic interactions. To attain this advanced level of control, a more thorough understanding of the interfacial interactions between biomolecules and materials is required. Undertaking both extensive experimental and computational studies, we have begun to unravel the influences and effects that drive biotic-abiotic interactions. In this talk, I will cover our approaches in understanding how biomolecules interact with abiotic surfaces, to control physiochemical properties by modulating these interactions, and in developing new routes for the synthesis and assembly of functional hybrid materials.

Although nanotechnology promises to revolutionize the prevention, detection, and treatment of countless diseases, the in vivo behavior of nanoparticles is extremely complex and, therefore, difficult to predict and control. Numerous variables have been reported to affect the biodistribution of nanoparticles, including size, shape, and surface charge. In order to advance our fundamental understanding of nanoparticle biodistribution, we developed mesoporous silica nanoparticle-supported lipid bilayers (‘protocells&’ - see Nature Materials (2011), 10: 389-397), which synergistically combine advantages of porous nanoparticles and liposomes and possess many features that can be independently and precisely modulated. We then used single-photon emission computed tomography (SPECT) and inductively-coupled plasma mass spectrometry (ICP-MS) to assess the influence of size, size distribution, shape, surface charge, surface modifications, dose, and route of administration on the time-dependent biodistribution of protocells in mice and rats. Our data indicate that size and route of administration have the most dramatic effect on biodistribution, followed closely by such properties of the protocell&’s supported lipid bilayer (SLB) as surface charge and conjugation with targeting and ‘self&’ ligands. We found that, upon intravenous (IV) injection, protocells with MSNP cores le; 100-nm in diameter and SLBs composed of zwitterionic lipids remained in circulation for up to 7 days, while protocells with MSNP cores > 150-nm in diameter and SLBs composed of anionic or cationic lipids rapidly accumulated in the liver and spleen. We also discovered that modifying protocells with CD47, a molecule recognized as ‘self&’ by innate immune cells, increased circulation times from 7 to 14 days, while modifying protocells with a single-chain antibody fragment (scFv) against aminopeptidase P, a molecule over-expressed by pulmonary vascular endothelial cells, triggered rapid accumulation in the lungs. Interestingly, we found no statistical correlation between circulation times and either protocell shape or dose. Specifically, protocells with spherical, 75-nm MSNP cores and protocells with rod-shaped MSNP cores with aspect ratios of 2, 3, and 4 remained in circulation for equivalent periods of time; biodistribution was also not affected by varying the IV-injected dose from 20 mg/kg to 200 mg/kg. Finally, we compared the biodistributions of 100-nm, zwitterionic protocells upon intraperitoneal (IP), intradermal (ID), oral, and inhalational (IH) administration and found that 42%, 17%, and 9% of IP, ID, and orally-administered protocells entered circulation within 4 hours, while 96% of IH-administered protocells remained in the lungs after 72 hours. Taken together, our results indicate that the time-dependent biodistribution of protocells can be entirely altered by varying their size and route of administration, as well as properties of the protocell&’s bio/nano interface (i.e. the SLB).

Chirality is a geometric property of objects that cannot be superimposed onto their mirror images. Chiral structures also give rise to unique chiroptical effects when interacting with polarized light. These properties are fundamental in biomolecular systems, and chiroptical spectroscopy is often used for their characterization. The role of chiral surfaces of inorganic crystals and their interaction with biomolecular systems has been discussed in many different theories that relate to the generation of homochirality in biomolecules. In this talk a somewhat complementary field will be discussed. In the first type of systems that will be described, induction of chiroptical effects in nanocrystals of achiral semiconductor materials using chiral biomolecules will be described.¹ This effect is interesting for fundamental studies of exciton - molecular level interactions, as well as exciton level structure characterization. However, it is generally very weak. More recently we introduced the concept of enantioselective synthesis of intrinsically chiral inorganic nanocrystals, which leads to more pronounced effects.² Many inorganic materials such as quartz, mercury sulfide and tellurium crystallize in chiral space groups with a chiral lattice. Biomolecules can be used to induce enantioselectivity in the nucleation and growth of nanocrystals of these materials. For the case of tellurium, we show that crystal growth in the presence of the small peptide, glutathione, results in nanocrystals where the atomic scale lattice chirality translates to the overall shape chirality on a 100 nm scale.³ This is a unique example for a colloidal chemistry approach for self assembling inorganic nanocrystals, which exhibit chirality at two size hierarchies. These systems may be useful for applications in metamaterials fabrication, asymmetric catalysis, sensing and optical devices. On a more fundamental level, these are excellent model systems for studies of chiral crystallization and separation, and the interaction of chiral biomolecules with chiral crystals. The possible role of chiral inorganic crystals and surfaces in the evolution of biomolecular homochirality has been considered by many researchers. Here it is demonstrated that the opposite effect, of biomolecules affecting chiral inorganic crystallization, is also intriguing.
1.Ben-Moshe, A.; Swarczman, D.; Markovich, G. " Size Dependence of Chiroptical Activity in Colloidal Semiconductor Quantum Dots" ACS Nano 5, 9034-9043 (2011)
2.Ben-Moshe, A.; Govorov, A. O.; Markovich, G. "Enantioselective Synthesis of Intrinsically Chiral Mercury Sulfide Nanocrystals" Angew. Chem. Int. Ed.52, 1275-1279 (2013)
3.Ben-Moshe, A.; Wolf, S. G.; Bar-Sadan, M.; Houben, L.; Fan, Z.; Govorov, A. O.; Markovich, G. "Enantioselective control of lattice and shape chirality in inorganic nanocrystals using chiral biomolecules" Nat. Comm. 5, 4302 (2014)

The interface between proteins and, extended or nanostructured, inorganic surfaces, is the key element of several current and potential biotechnological applications. For example, redox enzymes may be supported on electrodes, to create biofuel cells able to use fuel other than hydrogen [1]; peptides able to specifically bind a given surface have the potential to guide the self-assembling of nanosystems [2]. In a more biologically-oriented context, the interaction of amyloidogenic proteins with surfaces and nanoparticles is being investigated to understand how such interaction may affect the misfolding and fibrillation process [3].
Bare and functionalized gold surfaces, and gold nanoparticles, are particularly relevant in these frameworks. Au surfaces are often used for electrodes or to support self-assembled monolayers. Au nanoparticles are relatively cheap, can be synthesized with a good control of shape, size, and functionalization, and present useful optical properties.
We have developed and applied tools for the atomistic simulations of the interaction of proteins and peptides with various surfaces of gold [4,5], including functionalized ones, and with gold nanoparticles [6,7]. In this talks, such tools will be introduced and some examples of their applications relevant for material science (e.g., enzymatic biofuel cells [8]) and nanobiotechnology (e.g., interaction of gold nanoparticles with amyloidogenic proteins/peptides [7]) will be presented.
[1] Cracknell, J. A. et al. Chem. Rev. 108, 2439 (2008)
[2] Sarikaya, M., et al. Nature Mater. 3, 577 (2003)
[3] Linse, S., et al. PNAS 104, 8691 (2007)
[4] Iori, F. et al. J. Comp. Chem. 30, 1465 (2009)
[5] Wright, L. et al. JCTC 9, 1616 (2013); JPC C 117, 24292 (2013)
[6] Brancolini, G. et al. ACS Nano 6, 9863 (2012)
[7] Brancolini, G. et al. Nanoscale 6, 7903 (2014)
[8] Zanetti-Polzi, L. et al. JACS 136, 12929 (2014)

In this reaserch, we report the development of surfactant free-gold nanoparticle (AuNP)-DNA complexes that remained stable in solutions with extremely high ionic strength, using seawater as a model solution. Although the stability of AuNPs can be increased to a certain degree by functionalizing negatively charged DNA strands on their surface, they still have limited stability in highly concentrated salt solutions. However, we found that AuNPs functionalized with poly T (thymine) bases have exceptional stability in high ionic strength solutions. For example, AuNPs functionalized with a 5T spacer remained highly stable in seawater, with no color change and no red-shift in absorbance spectra for up to 9 days. Using this surprising property of poly T spacers, we prepared highly stable AuNP-DNA complexes containing random sequences by introducing 5T spacers on the random sequenced DNA strand. The random sequenced AuNP-DNA complexes remained stable in seawater, several molar concentrations of monovalent metal ion solutions (6.1 M Na⁺ or 4.8 M K⁺), and millimolar concentrations of diverse divalent metal ions. In addition, the highly stable AuNP-DNA complex maintained biological activity in seawater, which was demonstrated by complementary reaction and aptamer based biosensing. These results provide important insight into NP use for various applications under harsh biological and environmental conditions.

Adequate strength, tailored stiffness, and an interconnected porous architecture are crucial requirements for engineered bone tissue scaffolds; however high porosity reduces mechanical properties, making it difficult to fabricate scaffolds with the required mechanical properties (1). A potential solution to this on-going problem is the deposition of a stiff and strong polymer-nanocomposite coating onto a porous template using layer-by-layer (LbL) assembly. This technique involves the alternate deposition of oppositely charged electrolyte complexes (e.g. polymer and nanoparticles) onto a substrate resulting in the formation of multilayer films, and is one of the most versatile means to control film properties and molecular architecture at the nanoscale (2). The aim of this study was to adapt LbL assembly to fabricate nanocomposite coatings onto porous substrate, enabling customisation of mechanical properties and porosity to obtain materials suitable for bone tissue scaffold applications.
LbL deposition was conducted using aqueous solutions of polyethyleneimine (PEI), polyacrylic acid (PAA) and nanoclay (MTM) (Sigma Aldrich), prepared as previously described (3). Open-cell polyurethane (PU) foam substrates (30 PPI, EasyFoam Ltd.) were alternately subjected to cationic (PEI), and anionic (PAA and MTM) solutions using a custom-built apparatus. The deposition of a single PEI/PAA/PEI/MTM multilayer was repeated in intervals of 5 multilayers to obtain the desired number. The coated samples were dried for 24 h after each interval and the total mass of the coating was determined before deposition of subsequent multilayers. The mechanical properties of coated foams were determined by quasi-static mechanical testing in compression.
The physical properties of the multilayer film can be tailored by changing certain process parameters (4): pH of solutions, drying and deposition times, and salt concentration. These variables were systematically altered and the effect on deposition rate and mechanical properties were measured. The mass and thickness of the multilayer coating were found to correlate directly with the number of layers deposited. The elastic modulus of coated substrates increased by over an order of magnitude with the deposition of 60 multilayers. A micromechanical model for open-cell foams (5) was adapted to predict the porosity and mechanical properties of coated open-cell foams. It is expected that these results will serve as a guide to the design of scaffold materials with tailored stiffness and porosity within a suitable range for bone tissue scaffold materials.
References
1. Hutmacher, D. (et al.), J Tissue Eng Regen M, 1: 245-60, 2007
2. Yoo, D. (et al), Nano Lett, 8: 1762-70, 2008
3. Ziminska, M. (et al.), Proceedings of the 26th ESB Conference, Liverpool, UK, 2014
4. Dubas, S. (et al.), Macromolecules, 32: 8153-60, 1999
5. Ashby, M. (et al.), Phil. Trans. R. Soc. A, 364: 15-30, 2006

Consolidating physical science and engineering of surfaces with biology has produced novel nanomaterial interfaces which have the potential to make a major contribution to the field of clinical diagnostics. Nanoparticle-based multiplexing systems are particularly exciting as they can provide rapid, high-throughput analysis for very small sample volumes. Quantum dots (QDs), which have excellent potential for application here, are fluorescent semiconducting nanocrystals that have been used to produce a large variety of novel and highly sensitive assays [1]. However, there is still an unmet need for coatings and bioconjugations strategies in order to be able to use QDs both as a scaffold and in transduction of multiple biorecognitions events.
We have designed a novel multiplexing QD assay with the express intention of detecting and quantifying simultaneously both microRNA and protease activity in physiological conditions. The assay is based on a unique and systematic orthogonal peptide nucleic acid (PNA)/DNA duplex ‘tag&’ system that allows the detection of multiple analytes (in number and type). Additionally, the precisely engineered surface biofunctionalization of our QDs provides both high stabilization in physiological samples (urine, serum etc.), and specific conjugation groups for the attachment of the DNA ‘tags&’. The recognition of the target analytes is based on simultaneous binding between the unique DNA oligonucleotide on the surface of QDs and the complementary PNA on the detection complexes. These unique complexes are designed and synthesised as a linear combination of three components: i) a unique PNA tag, ii) detection substrate sequence (either a peptide for proteases, or a DNA sequence for miRNA or natural nucleic acids), and iii) a fluorescence energy acceptor (e.g. FRET dye, quencher). Then, the analytical signal will be correlated to the reduction of the fluorescent intensity of the QD due to the near presence of the FRET dye or quencher.
All together, these components yield a versatile detection system for various analytes in physiological conditions, and as such is an exciting development at the cutting edge of nanoparticle biosensing research.
[1. Howes, P.D., R. Chandrawati, and M.M. Stevens, Colloidal nanoparticles as advanced biological sensors. Science, 2014. 346(6205)]

Arrays of vertically aligned nanowires (NW) are increasingly being used as new nanometer-sized tools to probe biomolecular interactions¹ and manipulate living cells². In these cell-based applications, it can be anticipated that the topography of the NW arrays would have an effect on the plasma membrane as the NWs come in contact with the cell surface. In this study, we used phospholipid bilayers as models of cellular membranes and demonstrated that fluid supported bilayers can be formed on vertically aligned gallium phosphide nanowire forests via self-assembly from vesicles in solution.³ The dimensions of the nanowire arrays were designed such that the supported membranes on individual nanowires could be monitored in situ using fluorescence confocal microscopy. Using fluorescence recovery after photobleaching, we determined that the lipids are able to diffuse laterally within the NW-supported bilayer. The lipid bilayer is fluid and continuous, and its topography follows the contours of the surface formed by the array of NWs. Thus, as the vertical NWs have a very high aspect ratio, with nano-range diameters and micro-range lengths, there is a large bilayer surface available for the immobilization and study of biomolecules in a single field of view. We used the NW-supported bilayer to investigate the binding of membrane-associated proteins as well as the tethering of lipid vesicles to the bilayer. The ability to use bilayers to couple molecules of interest to the nanowire surface unlocks new possibilities for sensing of membrane processes.
References:¹ ten Siethoff, L., Lard, M., Generosi, J., Andersson, H. S. Linke, H. & Maring;nsson, A. Molecular Motor Propelled Filaments Reveal Light-Guiding in Nanowire Arrays for Enhanced Biosensing, Nano Letters, 2014, 14, 737- 742.; ² Piret, G., Perez, M.-T., Prinz, C. N. Neurite Outgrowth and Synaptophysin Expression of Postnatal CNS Neurons on GaP Nanowire Arrays in Long-Term Retinal Cell Culture Biomaterials, 2013 , 34, 875- 887.; ³ Dabkowska, A.P., Niman, C.S., Piret, G., Persson, H., Wacklin, H.P., Linke, H., Prinz, C.N. & Nylander, T. Fluid and highly curved model membranes on vertical nanowire arrays. Nano Letters, 2014, 14 (8), 4286-4292.

Metal nanoparticles (NPs) of various sizes and in particular of different shapes are highly interesting for their potential applications in the fields of biomedicine, sensors, metamaterials and light harvesting, due to their size and shape related optical (plasmonic) properties. They can be easily synthesized in large quantities and with high dimensional precision via wet chemical synthesis procedures. However, it is the coating material that ultimately limits the application of such metal NPs, due to the issues such as colloidal stability, wettability and toxicity, which strongly depend on the properties of the coating material. Nature has developed multi-functional polymers since the evolution of life, such as DNA/RNA, polysaccharides and proteins. Proteins are natural copolymers of polar, nonpolar, and ionic (anionic and cationic) monomers (in total 21 aminoacids) and are highly sensitive to temperature, pH, ionic strength, solvent, chirality and metal ions. Hence, protein coated nanoparticles provide access to a multi-functional and multi-responsive hybrid materials with controlled properties and bio-functions. In this work, we present protein capped metal nanoparticles [1-3] of various shapes, i.e. nanospheres and nanorods, which are extremely sensitive to pH, temperature and heavy metals, exhibiting a pronounced optical response, which can be monitored even by naked eye. Moreover, such NPs are extremely stable, mimicking somehow nature's concept of colloids and interfaces and consequently also highly biocompatible. Hence, such NPs exhibit remarkable physicochemical properties in biological fluids inside and outside living cells.
[1] M. Chanana, M.A. Correa-Duarte, and L.M. Liz-Marzán, Small, 2011, 7, 2650
[2] M. S. Strozyk, M. Chanana, I. Pastoriza-Santos, J. Pérez-Juste and L. M. Liz-Marzán, Adv. Funct. Mater., 2012, 22, 1436.
[3] M. Chanana, P. Rivera_Gil, M. A. Correa-Duarte, L. M. Liz-Marzán and W. J. Parak, Angew. Chem. Int. Ed., 2013, 52, 4179

Phosphate-based glasses (PBG) have wide application as biomaterials because they dissolve when implanted into the body, with a composition-dependent dissolution rate that varies over several orders of magnitude, and they can be synthesised containing therapeutically useful ions, antimicrobials or nutrients. This makes them potentially very useful as materials for targeted delivery of medical treatments.
In order for the therapy to be delivered at the appropriate rate, it is vital to understand the dependence of the glass dissolution on its structure and properties. We have made extensive use of classical and ab initio molecular dynamics simulations to look at the atomic-level structure of PBGs, beginning with the development of an accurate interatomic potential for phosphates [1]. The structures in the bulk glass which control the dissolution have been identified [2], as well as the effects on the structure of therapeutic dopants like fluorine [3], silver [4], or strontium [5].
However, glass dissolution occurs from the surface, and so the structure and properties of the glass surface will also be profoundly important. The structures (e.g. coordination numbers and electronic properties) on the surface will be very different to those on the bulk, which will effect the dissolution substantially. We present our ongoing work on understanding the surface environments of PBGs, including highlighting reactive sites and characterising the reaction of the glass surface with the body environment. Our ultimate aim is to understand the dissolution process at the atomic level, and optimise the glass dissolution rate for specific therapies, and we outline our progress toward that goal.
[1] R. I. Ainsworth, D. Di Tommaso, J. K. Christie, N. H. de Leeuw, J. Chem. Phys. 137, 234502 (2012)
[2] J. K. Christie, R. I. Ainsworth, D. Di Tommaso, N. H. de Leeuw, J. Phys. Chem. B 117, 10652 (2013)
[3] J. K. Christie, R. I. Ainsworth, N. H. de Leeuw, Biomaterials 35, 6164 (2014)
[4] R. I. Ainsworth, J. K. Christie, N. H. de Leeuw, Phys. Chem. Chem. Phys. 16, 21135 (2014)
[5] J. K. Christie, N. H. de Leeuw, in preparation

Developed biomaterials and their concept of controlled drug delivery system are proven their excellence in this research. The biomaterial, prepared using hydroxyapatite (HAp), shows a hollow structure with the presence of connections between inner and outer surface to load drug carriers. The poly (lactic-co-glycolic acid) nanoparticles as a drug carrier contain dexamethasone, which is known to cause osteoinduction. Surface of the drug carriers are modified using polyethyleneimine and therefore is able to conjugate to the surface of HAp granules. The hollow HAp granules, containing the drug carriers on inner and outer surface, show the controlled drug release rate compared to the granules, containing the drug carriers only on the outer surface. The pores that are designed for insertion of drug carriers and also the preosteoblast. Consequently, they influenced on the cellular behavior; the first is that the cell proliferation and the second is that the early stage of osteogenic differentiation. The effects of controlled release rate is evident up through the two weeks after the cell seeding, results in increase of osteogenic differentiation. From the above results, the drug carriers loaded hollow HAp granules are shown to be patient-specific biomaterials and exhibit the potential for hard tissue regeneration.

We have proposed a biological nanodevice fabrication process, which was named “Bio Nano Process” (BNP)¹⁾. Thre BNP produces nano-device key-components by biomineralization and self-organization of natural/artificial protein supramolecules. Protein supramolecules work as templates for the synthesis of homogenous nanoparticles (NPs) / nanowires (NWs) of metal-complexes and semiconductor materials ^2,3). The chemically or genetically modified outer surfaces makes them self-organize into functional nano-structures on a substrate. The addition of site specific biding peptides to the protein supramolecule is very powerful method to control both protein-protein and protein-substrate interactions. All process are carried out in aqueous solution, under ambient pressure and room temperature, which means that the BNP is a green process and environment-friendly.
We have already designed and produced several kinds of protein supramolecules for the BNP. Applying those, we have produced a floating nanodot gate memory (FNGM), a single electron transistor, a Re-RAM, bio-sensors, dye-sensitized solar cell, thermoelectric devices and other electronic devices ^2,4,5,6,7). Moreover, by combining the BNP and nano-etching, we fabricated arrays of silicon nanodisks⁸⁾ which were ideal quantum wells and are under investigation for quantum solar cell application. Proteins which have two types of site-specific binding abilities were genetically produced and the obtained proteins, “Proter Protein”, catch and deliver NPs / NWs to the specific patterns on a substrate⁹⁾. It is also expected that the BNP is well suited for plastic substrate devices.
We are now trying to expand the BNP frontier further. To realize such targets, it is true tha we need more exploration into the nano-space, where solid material surface and a cluster of biomaterials or proteins interfaces interact coordinately. We need how they interact, work coordinately there. We named the nano-space “Active Bio Field” and start expolataion. The combination of conventional technologies and the BNP are now producing and will produce nano-devices efficiently and economically, which could not be realized in their absence.
References 1) I. Yamashita, Thin Solid Films393, 12-18 (2001). 2) I. Yamashita et.al., Biochimica et Biophysica Acta1800, 846-857, (2010), 3) M. Kobayashi, et.al., Nano Lett., 10 (3), pp 773-776 (2010). 4) A. Miura, et.al., Jpn. J. Appl.Phys., 45, L1-L3, (2006). 5)S. Kumagai, et.al., Appl. Phys. Lett. 103, 223103, (2013). 6) I. Inoue et.al., ChemsusChem.7(10), 2805-2810, (2014). 7) M. Ito, et.al., Appl. Phys. Express7, 065102 (2014), 8) C. Huang et.al., Jpn. J. of Appl. Phys., 48, 04C187-1-04C187-6 (2009). 9) B. Zheng, et.al., Nanotechnology21, 045305, (2010)

Electrowetting on dielectric (EWOD), owing to its ability to electrically manipulate tiny individual droplets without involving movable mechanical parts, has received much attention in the past two decades [1, 2]. Despite tremendous promise, the use of solid dielectric layer between the aqueous droplet and underlying electrode is associated with inevitable physical and chemical heterogeneities [3, 4], leading to limited functionalities. For example, owing to the large contact angle (CA) hysteresis, contact line pinning [5] as well as CA saturation [6] at high voltage, it remains challenging to achieve reversible electrowetting with a large degree of switchability in ambient conditions. Moreover, activating droplet in EWOD is vulnerable to pronounced oscillation in response to an abrupt external stimulus, resulting in elongated time for the droplet to reach its equilibrium state [7]. Here, we demonstrate a new paradigm of electrowetting on bio-inspired soft liquid-infused film (EWOLF) that allows for the enhanced reversibility and faster response time to reach the steady state simultaneously. The liquid-infused film is achieved by locking a liquid lubricant in a porous membrane through the delicate control of wetting properties of the liquid and solid phases. Taking advantage of the negligible contact line pinning at the liquid-liquid interface [8, 9], the droplet response in EWOLF can be electrically addressed with enhanced degree of switchability and reversibility compared to the conventional EWOD. Moreover, we show that the infiltration of liquid lubricant phase in the porous membrane also efficiently enhances the viscous energy dissipation, suppressing the droplet oscillation and leading to fast response without sacrificing the desired electrowetting reversibility. Meanwhile, we find that the enhanced damping effect associated with the EWOLF can be tailored by manipulating the viscosity and thickness of liquid lubricant. We also demonstrate the feasibility of developing adaptive liquid lens for fast focusing using the as-proposed EWOLF.
References:
[1] B. Berge, C. R. Acad. Sci. II 317, 157 (1993).
[2] H. J. J. Verheijen and M. W. J. Prins, Langmuir 15, 6616 (1999).
[3] G. Manukyan, J. M. Oh, D. van den Ende, R. G. H. Lammertink, and F. Mugele, Phys. Rev. Lett. 106, 014501 (2011).
[4] G. McHale, C. V. Brown, M. I. Newton, G. G. Wells, and N. Sampara, Phys. Rev. Lett. 107, 186101 (2011).
[5] X. M. Chen, R. Y. Ma, J. T. Li, C. L. Hao, W. Guo, B. L. Luk, S. C. Li, S. H. Yao, and Z. K. Wang, Phys. Rev. Lett. 109, 116101 (2012).
[6] J. Liu, M. R. Wang, S. Chen, and M. O. Robbins, Phys. Rev. Lett. 108, 216101 (2012).
[7] S. R. Annapragada, S. Dash, S. V. Garimella, and J. Y. Murthy, Langmuir 27, 8198 (2011).
[8] A. Lafuma and D. Quéré, EPL 96, 56001 (2011).
[9] T. S. Wong, S. H. Kang, S. K. Y. Tang, E. J. Smythe, B. D. Hatton, A. Grinthal, and J. Aizenberg, Nature 477, 443 (2011).

In recent years, optical imaging using fluorescence emission in the second near-infrared window (NIR-II: 900 - 1,400 nm wavelengths) has emerged as a promising domain for in vivo sensing and detection. However, the application of these technologies for disease diagnostics has remained relatively challenging, due to the low availability of effective, target-specific molecular probes, and difficulties in harmonizing the interface between biological targets and functional nanomaterials. Here, we present applications of biologically-templated nanocomposites of M13 virus conjugated with single-walled carbon nanotubes (SWNTs) as probes for high-resolution noninvasive sensing and disease detection in living hosts. We have designed a highly customizable modular platform comprised of genetically engineered M13 bacteriophage as a biological scaffold playing the role of a multifunctional, targeted vector. This construct enables us to achieve a well-dispersed, stable aqueous suspension of wavelength-tunable nanoparticle fluorescent probes for optical imaging, as well as attach specific targeting agents to detect the disease of interest in vivo. Using a mouse model of ovarian cancer, we demonstrate that our peptide-labeled M13-SWNT probes can target tumors with high specificity and sensitivity, and assist in real-time surgical intervention, with sub-millimeter tumor resolution at a > 98% true positive rate. This is an order-of-magnitude enhancement compared to the current clinical gold standard, contrast-enhanced Computed Tomography, which has a detection sensitivity ~ 10% when the tumor diameter < 1 cm. In addition, using a mouse model of Staphylococcus Aureus endocarditis infections, we show our antibody-labeled M13-SWNT probe offers greater signal-to-background enhancement in deep tissue sensing of infectious diseases, achieved at an order-of-magnitude lower dosage compared to small molecule fluorescent probes. Our biotemplated nanomolecular probes offer distinct advantages of being (a) biologically functionalized without surface chemical modification of SWNTs, for retaining high photoluminescence; (b) aqueous dispersed, for in vivo applicability; (c) actively targeted, for highly specific detection; (d) modularly tunable, for detection of a wide range of diseases; and (e) low dosage achieving high contrast detection, for minimizing patient exposure. These results open up exciting new avenues for the exploration of detection and sensing technologies using M13-SWNT probes as a safer, non-ionizing (compared to X-ray CT), less expensive (compared to Magnetic Resonance Imaging) imaging modality. We aim to progress towards enhancing our sensing capabilities using a combination of custom-designed signal processing and sophisticated near real-time image analysis techniques for cellular level detection at tissue depths ~ 10 cm, thereby pushing the boundaries of optical imaging.

In the past decade and half, we have gained incredible insight into the workings of Nature at the smallest scales as we developed the ability to manipulate, control and interrogate matter at the atomic scale. But much of the promise of establishing new industry and economies founded on these scientific achievements has not come to pass. This is all about to change. With the foundations laid, the next 15 years is destined to see the application of these scientific advances into technologies that directly improve the human condition creating both economic and societal wealth.
The ability to use machines to manipulate matter a single molecule at a time renders many things possible that were impossible before. Living systems do this on a regular basis. The core challenge is how to transform a labile molecule that exists in a fragile living organism and to transfer that functionality into a stable system that is economically scalable. The most significant difficulties revolve around environmental stability and the inherent structural limitations of the molecule. Ingenuity Lab was created to bring together researchers from many disciplines to capitalize on the molecular interactions found in living systems and, through molecular manipulation, incorporate this functionality into complex systems to yield technology for solving many of the worlds societal challenges.
Presented is the concept of convergent technology, the intersection of the precision assembly of matter, nanotechnology, coupled with the functional building blocks of nature, biotechnology, and fused by the network flow of spatiotemporal information, informatics. We will present the details of the technological demands and the results of efforts associated with the production of a new class of functional material and devices. Elements of the discussion will include the genetic engineering of active biological molecules into engineering building blocks and the incorporation of “metabolism” into engineered devices and materials through precision assembly of these molecules into stable “active” materials. Finally, we will provide exemplars using these building blocks to engineer systems that address issues surrounding energy, environment and human health: the societal grand challenges of our age.

Nature abounds with mysterious biological creatures and organisms that exhibit unique surface wettability, such as lotus leaves with the self-cleaning property, rice leaves and butterfly wings with directional adhesion property, the mosquito eyes with antifogging functionality, and the Namib Desert beetle and spider silk with water collection ability. To obtain the artificial surfaces with unique wettability and functions, the combination of the unique structural designing of these biological materials or organisms with intrinsic material properties is crucial.
Among the special wetting properties, superhydrophobic surfaces with water repellent or water adhesive properties have received considerable attention as they have various applications including self-cleaning fabrics, anti-fog windows, drag reduction, and water transportation. In particular, in-situ manipulation of chemical or structural properties of superhydrophobic surfaces is greatly desired to control the motions of droplets for realization of advanced lab-on-a-chip devices for biological and chemical applications where minimal liquid-substrate interaction is required.
Here, we demonstrated a novel and facile method to control droplet motions on the polymeric superhydrophobic surface via the vacuum induced curvature formation. The superhydrophobic surface was composed of micro-pillar structures and the water droplets can be firmly adhered without deformation. When the vacuum pressure was locally applied to the superhydrophobic surface, the substrate was stretched and the water adhesion force of the mechanically deformed area was significantly decreased. The water adhesion force of the superhydrophobic surface can be easily adjusted and the motion of water droplets can be precisely controlled along the locally stretched area. Newer and emerging lab-on-a-chip applications using this droplet motion controllable superhydrophobic surface was demonstrated including stem cell spheroids formation, RNA transfection, and biomolecule analysis.

The understanding of the interface between living cells and artificial devices at the nanoscale is of crucial importance for advancing chip-based recording and stimulation techniques in neuroscience. Although chip-based devices such as microelectrode arrays (MEAs) are widely used and well-studied since decades, the emerging technology of 3D nanostructures for cell-chip coupling is currently a vivid field of investigation [1]. Our present study focuses on the investigation of cell-chip interfaces with optimized 3D nanoelectrodes for extracellular recordings. We have shown in a previous study that one cannot only effectively guide cells with mushroom-shaped 3D-nanoelectrode but also record electrical activity from electronic cells at the same time [2]. Additionally, we have investigated the principal deformation mechanisms of a bending membrane, which are caused by two kinds of nanoelectrodes: cylinders and mushroom-shaped structures. To this end, we have developed an artifact-free drying protocol of cells that enabled us to perform focused ion beam cross-section cuts through the cell chip interface and subsequent imaging with scanning electron microscopy [3]. Based on this previous approach, we have investigated a new resin-based embedding method, which drastically improves the image quality. The resin embedding maintains even all of the fragile inner cell components. By then performing carefully a cross-sectioning process, it is possible to determine the exact outline of the membrane deformation caused by the underlying nanoelectrode. Such membrane deformation measurements help to model better electric equivalent models of the seal resistances between the interface of cells and 3D nanoelectrodes. We investigated differently sized high and very high-aspect ratio nanostructures in cylindrical and mushrooms shape with respect to their biophysical behavior and electrical properties for cell-chip coupling. Here, we also introduce a completely new type of spine interface composed of thin stalks (stalk diameter: 150 nm - 350 nm) and extremely large mushroom caps (cap diameter: 2-4 µm). These geometrical features cause stronger engulfment of the nanoelectrodes by the cells and thereby improve extracellular recording capabilities. Such shape optimized nanoelectrodes are highly suited to achieve in-cell recording levels.
[1] Spira ME, Hai A. Multi-electrode array technologies for neuroscience and cardiology. Nat Nanotechnol. 2013 Feb;8(2):83-94. doi:10.1038/nnano.2012.265
[2] Santoro F, Schnitker J, Panaitov G, Offenhäusser A. On Chip Guidance and Recording of Cardiomyocytes with 3D Mushroom-Shaped Electrodes. Nano Lett. 2013 Nov 13;13(11):5379-84. doi:10.1021/nl402901y
[3] Santoro F, Dasgupta S, Schnitker J, Auth T, Neumann E, Panaitov G, Gompper G, Offenhäusser. Interfacing electrogenic cells with 3D nanoelectrodes: do position, shape and size matter. ACS Nano, 2014 Aug 8(7):6713-6723. doi:10.1021/nn500393p

Carbon nanotube (CNT) porins—short segments of CNTs inserted into a lipid bilayer membrane—are an ideal biomimetic system for studying ion transport in biologically relevant environments because the smooth, narrow, and hydrophobic inner pores of CNTs mimic natural biological pore channels. We will describe synthesis and optical properties of CNT porins and their assembly in lipid bilayer membranes. We will also report our measurements of osmotically-driven transport of charged and uncharged chemical species through CNT porins and regulation of the ion transport selectivity. These biomimetic membrane pore channels also open up ways to develop novel applications for biosensors, nanofluidic devices, molecular filtration, and artificial cells and cell-like structures.

Living organisms and biological substances are among the most difficult and persistent sources of surface fouling, particularly in medical and marine settings. The ability of organisms to adapt, move, cooperate, evolve on short timescales, and modify surfaces by secreting proteins and other molecules enables them to colonize even state-of-the-art antifouling coatings, and small surface defects can trigger protein aggregation and blood clotting. Attempts to combat these issues are further hindered by conflicting requirements at different size scales and across different species. Our recently developed concept of Slippery, Liquid-Infused Porous Surfaces (SLIPS) provides a defect-free, dynamic liquid interface that overcomes many of these problems at once. A single surface is able to prevent adhesion of a broad range of genetically diverse bacteria, including many pathogenic species that underlie widespread hospital-acquired infections, as well as marine algae. The same approach resists adhesion of proteins, cells, and blood, preventing clogging and thrombus formation inside medical tubing and catheters. At a larger scale, the slippery interface repels insects, barnacles and mussels, which slide off and actively avoid the coated surface. We are currently developing this strategy to solve longstanding fouling issues in a wide range of medical, marine, and other settings.

Biofuel cells are one of the most promising fields of research in sustainable energy. For biotechnological applications that split molecular hydrogen into electrons and protons, the oxygen-tolerant [NiFe]-hydrogenases are of particular interest [1]. In this regard, the immobilization of the membrane-bound-hydrogenase (MBH) of Ralstonia eutropha (R.e.) has gained significant interest in the last few years [1-3]. Fundamental studies are therefore required on the enzyme adsorption process, the elucidation of the electron transfer processes, the integrity and the activity of the immobilized enzyme which are crucial for understanding its functionality. The electrochemistry of R.e. MBH immobilized on graphite electrodes has been extensively studied [2][3], providing important information about its catalytic activity. Characterization of involved redox states as well as structure-function relationships have so far been mainly investigated in the bulk phase via Fourier-Transform-Infrared (FTIR) and Electron Paramagnetic Resonance (EPR) [4], however gaps exist which prevent the complete elucidation of the R.e. MBH functionality. In order to understand the correlation between the immobilization processes and the catalytic behavior of the surface immobilized R.e. MBH, elementary studies have been carried out by a spectroelectrochemical method, coupling direct enzyme film voltammetry and Surface Enhanced Infrared (SEIRA) spectroscopy. In this respect, influencing factors on the immobilization process and enzyme orientation including surface functionality, surface charge, pH dependence and Ionic strength will be discussed and compared to molecular dynamic (MD) simulations.
[1] B. Friedrich, J. Fritsch, and O. Lenz, “Oxygen-tolerant hydrogenases in hydrogen-based technologies,” Current opinion in
biotechnology 3 (2011) 358-64.
[2] J. Cracknell, A. F. Wait, O. Lenz, B. Friedrich, and F. a Armstrong, “A kinetic and thermodynamic understanding of O2 tolerance in
[NiFe]-hydrogenases.,” PNAS 49 (2009) 20681-6.
[3] F. Armstrong and J. Hirst, “Reversibility and efficiency in electrocatalytic energy conversion and lessons from enzymes,” PNAS 34
(2011) 14049-54.
[4] M. Saggu, I. Zebger, M. Ludwig, O. Lenz, B. Friedrich, P. Hildebrandt and F. Lendzian, “Spectroscopic insights into the oxygen
tolerant membrane-associated [NiFe] hydrogenase of Ralstonia eutropha H16.,” JBC 24 (2009) 16264-76.

Predictably interfacing biological molecules with solid materials is the key for drug delivery, enzyme immobilization, biofunctionalization of implants, and signal transduction in biosensors. Highly specific interactions controlled by proteins enable explicit recognition of minerals and formation of intricate supramolecular architectures in nature. Mimicking natural proteins, engineered short polypeptides have recently become ubiquitous molecular tools in the addressable functionalization of and organizations at solid interfaces. Furthermore, the simplicity of their short amino acid sequences and the presence of functional domains offer the potential for tailoring and interrogating individual types of intermolecular forces through rational mutation and design. Direct experimental observation of the interaction of such peptides with solids requires well-defined solid surfaces (i.e. atomic-scale topography, crystal structure, or surface chemistry) that are kept persistent under biological conditions, e.g., pH and buffer. These requirements have recently been realized with 2D solids, such as layered dichalcogenides, nitrides, and carbides, and these systems have been used to characterize the adsorption and assembly of solid-binding peptides. Here, using graphite-binding peptides originally selected by a phage display library, we demonstrate control of peptide-peptide intermolecular forces as well as peptide-solid interactions through rational mutation of their chemical and structural domains. The fundamental knowledge of bio/nano interfacial interactions is the key to controlling these molecular forces, leading to many self-assembled peptide (SAP) biomolecular surface structures. Proof-of-principle examples of technological implementations include applications in biosensing and biomineralization.

The contact of two hydrophobic surfaces in water is of importance in biology, catalysis, material science, and geology. A tenet of hydrophobic attraction is the release of an ordered water layer, leading to a dry contact between two hydrophobic surfaces. Although the water-free contact has been inferred from numerous experimental and theoretical studies, this has not been directly measured. I will present the use of surface sensitive sum frequency generation spectroscopy to directly probe the contact interface between hydrophobic surfaces Even though both these surfaces are identical in dry contact they differ dramatically in wet contact. The consequences of water trapped between these two hydrophobic surfaces will be discussed in context with adhesion and wetting.

Natural particles, such as pollen, diatoms, and fungal spores, exhibit a remarkable breadth of complex solid surface ornamentations as well as a nanoscale thin liquid glues. This talk will examine pollen, and show that these features combine to yield a natural pressure-sensitive adhesive system. In particular, we have discovered that the combination of solid spines or reticulate structures with the nanometric liquid pollenkitt gives pollen a load-sensitive adhesion on rough surfaces. A second discovery is that the pollen adhesion can be optimized when the adhesion surface patterns form a negative impression of the pollen spine features. These results are derived from measurements of pollen adhesion and detachment from surfaces with controlled roughness and pattern regularity, by using AFM and centrifuge methods. Three pollen species were investigated, each having a unique surface morphology and pollenkitt volume. Surface patterning of the test substrate was controlled by using blends of poly(styrene) with poly(styrene-b-isoprene) copolymer. Significant enhancement in the adhesion was observed for pollen deposited on rough patterned surfaces owing to multiple spine interactions with a rough surface. The adhesion was optimized when the counter surface pattern matched the spacing of the pollen spines. Modeling of pollen detachment behavior under centrifugal forces shows that the mechanism of pollen detachment switches from sliding to rolling as roughness increases. The pollen adhesion system provides a natural model for development of novel pressure-sensitive adhesives on rough or patterned surfaces.

The role of water molecules on the adhesion property of cellulose/phenolic adhesive interface is investigated by molecular dynamics simulations. Cellulose is one of the most abundant substances in the world, and the major constituent in the wood structure. Phenolic adhesive is largely used in the wood manufacture for gluing the wood panels together. The cellulose/phenolic adhesive interface is a representative of the interface between the wood panels and adhesives in the wood products. As the wood panels and adhesive are sensitive to environmental humidity, the interfacial adhesion of such interface when subjected to a humid environment can be a major factor in the durability of final products. Here, the simulation results reveal that water molecules significantly reduce the adhesion energy between cellulose and phenolic adhesive of 86.5%. Meanwhile, it is demonstrated that the adhesion energy can be recovered after the interface experiences further dry conditioning. The hydrogen bonds between the cellulose and phenolic adhesive are found to account for the strong interfacial adhesion, which can be interrupted in the presence of water molecules and recovered after further dry conditioning. The durability of the wood products is mainly determined by water molecules absorbed at the bilayer interface, which should be considered in a wet condition.

Over the past two decades, many bioadhesives including acrylates and fibrins gained significant interest due to their several advantages. Still at present, a universal bioadhesive that fulfils all the requirements such as the biodegradability, biocompatibility, controlled on demand adhesion, and adequate mechanical properties does not exist. Fine tuning of mechanical and drug-delivering properties of polyesters makes them a natural choice towards tissue fixation and controlled drug delivery applications.
Here, we report the development of polyester thin films modified with novel bioadhesive chemistries for “on-demand” tissue adhesion and local drug delivery.
The synthesis and functionalized films were characterized using 1H and 13C-NMR. The kinetics and mechanism for on-demand free radical adhesion to tissue were investigated using UV spectroscopy. Ex-vivo swine aorta tissue bioadhesion was assessed for the photosensitive films towards shear force.
The reaction kinetics study demonstrated that half-life of newly synthesized moiety was found to be 11.5 ± 0.9 min at 0.5 ± 0.1 mW/cm2 UV light. The reaction followed first order kinetics. Ex-vivo swine aorta tissue bioadhesion was found to have highest bioadhesion force, i.e. 450 ± 50 mN cm-2 compared to untreated films (p<0.05).

We are engineering oligonucleotide nanostructures that interact with non-nucleic acid macromolecules to achieve technologically useful purposes. We will illustrate this with 3 examples:
(1) Using SWNTs labeled with non-covalently attached DNA linkers are positioned on complementary DNA hooks presented by DNA origami. This allows assembly of cross-junction transistors by mixing origami templates with two types of labeled nanotubes.
(2) Engineering DNA linkers that assemble solution dispersed SWNTs into parallel arrays via surface diffusion. Duplex portions of the DNA linkers act as spacers to precisely control the inter-nanotube separation distance down to < 3 nm, and can serve as scaffolds to position additional components such as proteins between adjacent parallel nanotubes.
(3) Developing programmable chimeric oligonucleotide logic gate that convert RNA inputs of one sequence into RNAi outputs targeting an entirely different sequence. This signal gated RNAi molecule achieves high ON:OFF ratios of RNAi activity in human cells and opens potential new avenues for targeted molecular therapy.
We will present experimental and simulation results that demonstrate functionality of these systems, elucidate their crucial molecular scale features, and suggest steps for further optimization

The physiochemical properties (large surface area, extreme conductivity, high mechanical flexibility, and superior thermal transport) of graphene have attracted significant attention as a 2-D material for high performance electronics. Due to these unique properties, graphene is an attractive transducer for biosensing applications. However, the development of graphene-based sensors is still in its infancy needing further development. The understanding of the interfacial interactions between the biotic (biomolecules) and abiotic (graphene) component is critical to design an effective sensor. Similar to other transducers, bio-recognition elements (BREs), such as antibody, proteins, DNAs, and other small molecules are interfaced with the graphene to enable selective capture of target molecules. Peptides that were previously identified to be binding selectively to graphene are further investigated to describe how factors such as graphene quality and number of graphene layers, along with supporting substrate, influence the graphene-biomolecular interactions.
Distribution Statement A. Approved for public release, distribution is unlimited.

The accumulation of biomass on marine structures (such as ship hulls) is a major problem for maritime operations as it is associated with increased fuel consumption, damage and corrosion as well as the spread of invasive species. With commonly applied biocidal antifouling coatings coming under more and more legislative pressure, viable alternatives are needed. Here we present an effective, non-toxic, slippery surface technology for marine fouling prevention applications, demonstrating promising performance in laboratory and field-based marine fouling studies.

Surface Plasmon Resonance (SPR) has been used few decades for label-free detection and characterization of biochemical kinetics and affinities for many different types of analysts. The physical phenomena is not limited to biochemistry, but is applicable to many other nanoscale characterization.[1]
Aside of the traditional interactions, Multiparametric Surface Plasmon Resonance (MP-SPR) can be utilized to determine unique refractive index (RI) and thickness (d) of ultrathin films (d 0.5-100 nm) without knowledge of the RI of the material. These are important properties for many thin film coating industries and applications, and important knowledge in biomaterials also. The new method allows measurement of these properties for both dielectric layers, but also for metals and metal-like coatings that are difficult to measure with other optical methods.
Two new methods utilizing MP-SPR have recently been introduced, either measuring in two different media (2M) with high RI difference, such as air and water [1-3], or at two or more different wavelengths (2W) of light [2,3] in order to characterize properties of ultrathin films.
Strongly light absorbing materials, such as metals and semiconductors, have been difficult to measure with traditional thin film measuring optical methods, such as ellipsometry because they are mostly non-transparent materials [4]. For plasmon generation needed for MP-SPR this is not as crucial issue and even multilayered nanolaminates can be characterized.
With the ability to characterize both kinetics and nanoscale layer properties, MP-SPR is an effective tool for nanomaterial, biomaterial and biochemical interactions research. This makes the MP-SPR a powerful tool for multidisciplinary research, where both material physical- and interaction properties are characterized.
[1] Albers, Vikholm-Lundin, Nano-Bio-Sensing, 1st, Springer 2010
[2] Liang, et al., Sens.Act.B,149(1), 2010, 212-220
[3] Granqvist et al. Langmuir 29 (27), 2013, 8561-8571
[4] Hilfiker et al., Thin.Sol.Films, 516, 2008, 7979-7989
[5] Sadowski et al. Opt.Eng., 34 (9), 1995, 2581-2586

Nanomaterials such as nanowires and nanopillars have been shown to interact strongly with biological cells. Such 1D nanomaterials can enable the penetration of cell membrane by electroporation which is useful in the area of molecular biology to deliver polar substances into cells. Here we introduce a new application, water disinfection, enabled by 1D nanomaterial-assisted electroporation. The high electric field induced by the sharp 1D nanomaterials can be used to damage the membrane of the microorganisms and inactivate the microorganisms by disrupting the inner cellular environment. The 1D nanowires can enhance the local electric field 2-3 orders of magnitudes higher than that of planer structure. Using flow devices with filter electrodes made from silver nanowires or copper oxide nanowires, we demonstrate a high efficiency inactivation of both model bacteria and virus, of > 6log (>99.9999%) removal, with a fast treatment speed of 3000-15000 L/h-m², which is equivalent to only 1s of contact time of microorganism with filter electrodes. During operation, the flow device can be powered by a small voltage < 20V and the energy consumption was very low of only <100J/L. The material release to effluent water was shown to be minimal and within drinking water standard. This 1D-NE showed great potential as an energy efficient and low-cost alternative to chlorine and UV disinfection in water treatment applications.

Exosomes are submicron vesiclar particles secreated from almost all cells. Exosomes carry the information of the host cells that released them and circulate in bodily fluids including blood, urea saliva etc., and have been attracting attentions for their potential use in diagnosis and therapy because of their possible involvements in the development and metastasis of various maladies. Here, we purified exosomes from various cancer cell lines using the density gradient centrifugation, incubated them with various inorganic materials and then characterized their adsorption behaviors by AFM. Sizes, shapes and amounts of the adsorbed exosomes varied depending on the substrates as well as sources of cancer cell lines used. For the development of diagnostic and therapeutic devices for exosome-based medicine, basic knowledge of the interaction between exosomes and the surfaces of various materials is absolutely vital.

Many active biomolecules are available to perturb cell function by direct addition to cell culture media, including receptor ligands and antibodies. Drug delivery mechanisms interact with the cell membrane to deliver molecules directly into cells and generate exponentially more possibilities for cell perturbation, especially the option for effecting permanent change in cells by altering their genome, and the ability to directly perturb the central dogma by altering mRNA transcription and translation into protein. The majority of these active biomolecules have similar molecular structure, which led to the development of delivery vehicles specific for delivering DNA, peptides, and other classes of molecules. Recently, synthetic biomolecules have been developed which have hybrid characteristics, often due to the addition of functionality such as fluorescence or tailored binding sites. These synthetic functionalities could represent another exponential leap forward for studying cells in theory, but these molecules often lack an option for intracellular delivery due to their hybrid structure, limiting their practical use. Here we show the use of a microfabricated supported nanotube structure, called nanostraws, to deliver synthetic biomolecules into cells. The nanostraws are a nanobio interface, which create fluidic conduits into the cell for molecular transport. We focus on two types of molecules: synthetic carbohydrates with click chemistry moieties that, when delivered into cells, can be used to track metabolic pathways, and modified peptides with active sites that can be used to track enzyme activity. The delivery of these disparate molecules using the same technique demonstrates the utility of nanostraws as a non-specific delivery mechanism, capable of delivering synthetic molecules of diverse structures.

Viruses can be engineered to exploit validated genetic pathways known to be deregulated in many cancers. One limitation of oncolytic viruses for clinical applications is that they get cleared from the ciruclation after they get administered. To overcome an immune response and to enhance its potential use to treat primary and metastatic tumors, a method for liposomal encapsulation of adenovirus has been developed. The encapsulation of adenovirus in non-toxic anionic lecithin-cholesterol-PEG liposomes ranging from 140-180nm in diameter have been prepared by self-assembly around the viral capsid. The encapsulated viruses retain their ability to infect cancer cells. Furthermore, an immunoprecipitation (IP) technique has shown to be a fast and effective method to extract non-encapsulated viruses and homogenize the liposomes remaining in solution. 78% of adenovirus plaque forming units were encapsulated and retained infectivity after IP processing. Additionally, encapsulated viruses have shown enhanced transfection efficiency up to 4X higher compared to non-encapsulated Ads. The developed encapsulation design has the potential to enhance oncolytic viral gene therapy.

Recent research in tissue engineering and regenerative medicine is increasingly revolving around effective fabrication techniques to create functional scaffolds for cell manipulations. Specifically, three-dimensional (3D) supports presenting temporal and spatial control over the exposure of protein cues are desirable as they would allow spatial control over cell behavior [1-2].
Thus, in order to mimick the compositional diversity of natural extra-cellular matrices (ECMs) we propose here a new and versatile method to obtain synthetic ECMs, which are potentially directly applicable to clinical practices. Fabrication of three-dimensional (3D) gradients of proteins within microporous, biodegradable tissue engineering scaffolds making use of solution wetting is described. 3D regularly layered starting supports are manufactured by rapid prototyping of poly-ε-caprolactone (PCL). Uniform coating of the scaffold surfaces with “grafted-from”, poly(ethylene gycol)(PEG)-containing polymer brushes is employed for the covalent immobilization of proteins. The 3D gradient formation processes make use of surface energy and capillary forces, which administer proteins from their solutions inside the pores. Following these approaches multidirectional gradients of different protein species can be produced with precise control over the protein coverage morphology. In addition, 3D gradients of brush-supported fibronectin permit the controlled immobilization of human mesenchymal stem cells (hMSCs) in spatially determined cultures.
PCL-POEGMA scaffolds with 3D protein gradients demontrate to be easily reproduced also in clinics since they do not require any inert environment or complicated chemistry. Brush-coated scaffolds could be simply incubated with the required proteins and directly applied to patients in or without the presence of cell preparations. In this context, the versatility of this technique support the combination of multiple cues which could eventually trigger cell adhesion, migration or differentiation, in a single 3D porous structure. All the above-mentioned features make the method presented a practical and affordable strategy to synthetically mimic natural ECMs and their 3D multidirectional diversity.
1. M. P. Lutolf et al. Nat. Biotech., 2005, 23, 47-55.
2. J. Genzer, Ann. Rev.Mat. Res., 2012, 42, 435-468.

In this presentation we describe attempts to develop a range of bone- and nacre-like structural materials using freeze-casting technique, which utilizes the intricate structure of ice to create hybrid materials with complex lamellar and/or mortar and brick structures modeled across several length-scales. The challenge is to develop a new generation of implant materials that will combine the advantages of ceramics, in particular their inertness, with a mechanical response comparable to those of implant alloys. The unique properties of ceramic materials, including their outstanding corrosion resistance and excellent aesthetics, make them appealing candidates for many implant applications. However, their inferior mechanical properties, particularly fracture toughness, until now have hampered their commercial use, especially in load-bearing situations. This is a particular problem when designing porous implants or implants with rough and porous surfaces for better osseointegration. While the porosity will improve osseointegration, increased porosity can easily result in diminished strength and fracture at relatively low loads. Through new fabrication technologies, these limitations may be overcome. Our results show ceramic-polymer hybrid materials with toughness well in excess of those expected from a rule of mixtures construction. The architecture and properties of the synthetic materials are compared to their natural counterparts in order to identify the mechanisms that control mechanical behavior over multiple dimensions and propose new design concepts to guide the synthesis of hybrid/hierarchical structural materials with unique mechanical responses. The ultimate goal is to produce materials and therapies that will bring state-of-the-art technology to the bedside and improve quality of life and current standards of care. Work supported by the National Institute of Health under grant number NIH/NIDCR 1 R01 DE015633.

While nanotechnology has a number of biological applications, the lack of high-throughput, customizable approaches limits its scalability to biotechnology. Here, we address this limitation by developing a novel approach for rapid and inexpensive fabrication of polymeric micropatterned nanowire arrays. We describe two variations of this approach that allow for fabrication of both flat nanowire arrays and detachable nanowire-coated microstructures with tunable nanowire dimensions and go on to investigate biological applications for the resulting polymeric structures. First, we demonstrate that the micropatterned arrays of densely packed nanowires facilitate high-throughput, low-waste drug and reagent loading with micron-scale resolution via capillary action. Second, we show that micropatterned nanowire arrays provide hierarchical cellular control by directing cell shape on the micron scale and influencing focal adhesion formation on the nanoscale to provide a phenotype not achieved through micro- or nanotopography alone. This high-throughput nanofabrication approach has a number of possible applications in scaffold-based cellular control, biological assay miniaturization, and biomedical microdevice technology.

Besides the biological growth factors, small organic molecules, and chemical ions, physical signals is the other category of very important factors to tune/regulate the fate of stem cells. Recent years, more attention has been paid on the differentiation of stem cells on the physical signal, including, electric or magnetic field, surface topology of biomaterials, photo irradiation, and even pressure and strain from the materials. With progress of research in this field, some cures of connection between physical signal and bio pathway for differentiation have been discovered. However, more phenomena have still not been understood. Because the physical signals possess controllability and can be localized in a specific area, they are benefit to be used in tissue engineering for tissue regeneration. Therefore, finding new physical approaches for regulation fate of stem cells is a great challenge for alive biomaterials design and applications.
Recent year, some novel phenomena about the effect of physical signal on stem cell differential has been noticed. For example, nano-network morphology of HAP film can differentiate bone marrow mesenchymal stem cells (MSCs) to vascular endothelial cells, surface charges on LiNbO3 wafer can regulate MSCs differentiate to osteogenic cells, and a pressure from biomaterials can differentiate MSCs to neural cells.
In this talk, we will report the above works, and try to explain the reasons for physical signal induced differentiation from both physical mechanism and bio pathways. We believe that the regulation effect of physical signal will attract more attention, and will have great impact for design and application of biomaterials, especially for tissue engineering scaffold, and will bring great progress in tissue regeneration medicine.

DNA-based electrochemical sensors can be defined as nucleic acid layers with electrochemical transducers. DNA is especially appropriate for biosensing applications because interactions between complementary sequences are specific and robust to provide a simple and accurate biosensor for patient diagnosis. Electrochemical methods are appropriate for DNA diagnostics, giving a direct electronic signal [1]. But, there is still an extensive discussion about how charge transport occurs over DNA distance.
MicroRNAs are small sequences that regulate a wide range of cellular processes. There has been a huge interest in studying their expression in human cancers [2]. We proposed to transform MicroRNA into DNA and develop a “MicroDNA” electrochemical biosensor to detect miR-200a sequence (22-mer) related with breast cancer.
Au electrodes were cleaned using standard procedures [3] and experiments were performed at room temperature. 1 µM thiol - modified single-stranded probe sequence was dissolved in a pH 7 DNA solution (1 M phosphate buffer, 1 M NaCl, 5 mM MgCl₂ and 1 mM EDTA), and immobilized for 18 h. Sequence 1 mM 6 - mercapto - 1 - hexanthiol in MilliQ water was immobilized for 1 h and the hybridization with complementary strand dissolved in pH 7.4 hybridized solution (10 mM phosphate buffer, 1 mM EDTA, 1 M NaCl) for 1h.
Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were applied to control biosensor development. CV was carried out in a 10 mL electrochemical cell composed by modified working electrode, Ag|AgCl|1 M KCl reference electrode and platinum wire as auxiliary electrode. The measurements were performed in 5 mM potassium ferrocyanide and 10 mM phosphate buffer, cycled from -0.1 to 0.5 mV at 100 mV.s^-1. EIS was performed in the same electrochemical cell used for CV. Frequencies varied from 100 kHz down to 0.05 Hz with 10 mV amplitude and single sine.
The solutions employed in each step were studied and proved to be crucial to overall device performance. Specially MgCl₂ in DNA immobilization solution neutralizes the negative charges of single-stranded probes and promotes a huge surface coverage. The increase of immobilized single-stranded improves the electrochemical impedance spectroscopy hybridization signal. The analytical curve indicates the percent of hybridization as a function of complementary strand concentration. The results of electrochemical techniques will be also compared to quartz crystal microbalance experiments.
This work was funded by CNPq, CAPES and FAPESP Brazilian agencies.
Reference
[1] T. G. Drummond, M. G. Hill and J. K. Barton, Electrochemical DNA sensors. Nature Biotechnology 21(2003) 1192 - 1199.
[2] A. Esquela-Kerscher and F. J. Slack, Oncomirs - microRNAs with a role in cancer. Nat. Rev. Cancer 6 (2006) 259—269.
[3]L. M. Fiscger, M. Tenje, A. R. Heiskasen, et al. Gold cleanning methods for electrochemical detection applications. Microelectronic Engineering 86 (2009) 1282 - 1285.

In natural and anthropogenic environments, bacteria form interface-associated, heterogeneous communities called biofilms. These communities often comprise multiple species, and their stability and function are determined through interspecies interactions via specific chemical signaling or metabolite exchange. Moreover, cell phenotypes and community function are sensitive to local perturbations in environmental conditions. Here, we develop materials to engineer the structure and function of coculture biofilms whereby growth substrate topography induces switching in a signaling pathway between a commensal Escherichia coli strain and an opportunistic pathogen, Pseudomonas aeruginosa. The morphology of biofilms of E. coli on periodically patterned surfaces changes with the aspect ratio of nanostructures on the growth substrate. These morphological changes result in the accumulation of a metabolite which allows E. coli biofilms to resist displacement by P. aeruginosa, providing a molecular mechanism for tuning interspecies interactions. The connection between substrate structure and biological signaling allows us to deterministically engineer the composition and properties of these coculture communities. In this manner, we also demonstrate how growth substrate topography alone is capable of cultivating probiotic biofilms which exhibit long-lived resistance to pathogen colonization and are susceptible to antibiotic treatment.

Colloidal semiconductor nanocrystals are under intense development for a range of applications, including light-emitting devices for energy-efficient displays and light sources, light-harvesting elements for exploitable and low-cost photovoltaics, logical elements for quantum computing, as well as for many biomedical applications such as imaging, drug delivery, and sensors. In particular, the key advantage of using of artificial nanocrystals in biology, medicine and pharmacology is an ability to manipulate the units of matter at the scale close to molecular one. At this scale many biological processes involve mechanism of molecular recognition, and it is well known that chirality plays a key role in this mechanism. Potentially any nanocrystal can be intrinsically chiral due to their low symmetry and presence of bulk and surface defects. Chirality of the semiconductor nanocrystals makes possible their laquo;lock and keyraquo; interaction with biological objects since nanocrystals&’ sizes are comparable to the sizes of biomolecules and the pores of cell membranes.
Here we investigate intrinsic chirality of CdSe nanocrystals using a technique of circular dichroism (CD). We develop an experiment under assumption that as-prepared ensemble is a mixture of achiral nanocrystals and equal amounts of nanocrystals with opposite chirality, and possesses no optical activity. Consequently, to obtain an experimental evidence of intrinsic chirality of the semiconductor nanocrystals, it is necessary to separate their enantiomers. We show that optically active the quantum dots and quantum rods samples can be obtained by preferentially extracting either L- or D-enantiomers of nanocrystals from optically inactive sample. We use an original method of the enantioselective phase transfer of the nanocrystals from organic solvent to water assisted by the chiral molecules of ligand. This method allows us to isolate L- and D-enantiomers of the nanocrystals into different organic and aqueous phases and investigate their CD behaviour. The aqueous phase is enriched with nanocrystals with the same chirality as the ligand molecules used for the phase transfer, for example, L-nanocrystals in the case of using L-cysteine. In the contrast, the organic phase is enriched with the nanocrystals, whose interaction with chiral ligands was ineffective. These nanocrystals are still capped by achiral ligands. The aqueous and organic fractions possess almost mirror image CD signals, but the value of intrinsic CD signal in organic solvent is less. The reason for this difference may be in induced increasing of CD after adsorption of chiral ligands. The experiments with other types of quantum-confined objects such as nanoplatelets, nanotetrapods, and dot-in-rods also show increasing of CD signal after capping with chiral ligands. Importantly, CD increasing differs for different types of nanocrystals. In particular, CD increasing up to 30 times is observed for the nanoplatelets.

A new concept of organic-inorganic interface stabilization via specifically selected peptide-polymer conjugates can be successfully applied for the stabilization of nanoparticles and the improvement of mechanical properties of hybrid materials. The concept is based on the fact of the sequence specific recognition of the inorganic surface with peptides.¹ The idea of compatibilizer in form of peptide- polymer conjugate is inspired by nature examples like nacre or bone, where proteins or conjugated systems (proteoglycans) are able to recognize the inorganic surface and to implement the inorganic component into organic matrix.² Besides that compatibilizers seem to play an important role in the outstanding mechanical performance of biomaterials.
The compatibilizer combines a selected peptide sequence, specifically adhering to inorganic surfaces, and a polymer-block to compatibilize the inorganics with the polymer matrix. Peptide sequence was biocombinatorially selected from a phage display library containing ~10⁹ different sequences.³
Peptide-PEO conjugate as a tailor-made compatibilizers is incorporated in the polymer composite material, composed of MgF₂ submicron particles and PEO, which is considered as a model system for complex biomaterials. Filling of a polymer with inorganic particles normally leads to higher elastic moduli and the corresponding higher stiffness. Addition of the conjugates to the system leads to increased stiffness of material. So new, compatibilized composites with just 10 wt% MgF₂ of filling reach the same elastic modulus as a non-compatibilized composite with 40 wt% MgF₂. Change of the sequence of amino acids in the peptide does not lead to improved mechanical properties, indicating specificity of compatibilizer and peptide-particle interaction. Moreover, the fracture mechanism upon deformation is changed. The material structure is studied with electron microscopy techniques (SEM, TEM) and AFM. Interaction of peptide sequence with the inorganic surface is studied with NMR in the solution. The interactions in material and the changes in the material are followed with IR spectroscopy. The new concept of interface stabilization and resulting polymer-based materials might enable to draw links to biological systems like for instance bone and offer the new ways for the design of biomimetic materials.
¹T. Schwemmer JACS, 2012, 134 , 2385-2391.
²H.G.Börner, Prog. Polym. Sci. 2009, 34, 811-851.
³F.Hanszlig;ke, Adv. Mater. 2014, submitted

It is well known that layered host compounds intercalate various ions and molecules, which brings about the interlayer expansion. Recently we found that layered metal oxides undergo enormous and instantaneous swelling upon contact with a dilute solution of amines or organoammonium salts [1]. We observed that the sample volume dramatically expanded and, under the optical microscope, platelet crystals of layered oxides greatly elongated into a string-like object in a few seconds when brought into contact with the solution. The swollen crystals reverted mostly to an original size and shape immediately upon dropwise addition of an acid solution. SAXS measurements indicated the interlayer expansion to ~90 nm, corresponding to ~100 times the original spacing. Such a largely expanded interlayer space was predominantly occupied by water (>97 wt%) together with a trace amount of amine. Systematic studies [2] demonstrated that the degree of swelling is virtually independent of amine identity but is dominated by the concentration. On the other hand, the amine varieties affect the stability of the swollen structure; small and polar amines tend to produce the stable swelling while larger amines with a symmetrical molecular shape lead to exfoliation [3]. The results provide important knowledge for high-quality production of 2D molecularly thin oxides, and also may shed lights to peculiar behaviors of water confined in a restricted space.
[1] F. Geng, R. Ma, A. Nakamura, K. Akatsuka, Y. Ebina, Y. Yamauchi, N. Miyamoto, and T. Sasaki, Nat. Commun. 4 (2013) 1632.
[2] F. Geng, R. Ma, Y. Ebina, Y. Yamauchi, N. Miyamoto, and T. Sasaki, J. Am. Chem. Soc. 136 (2014) 5491.
[3] R. Ma and T. Sasaki, Adv. Mater. 22, 5082 (2010).

Biosensor technologies require the understanding of interactions between organic and inorganic materials to tune electric response functions. Peptide assembly on well-defined multilayer graphene substrates exhibit high biological selectivity and sensitivity towards analytes. Laboratory characterization of specific interactions and molecular assembly of such biomolecules on graphene surfaces in atomic resolution remains challenging and can be complemented by molecular simulations, as well as by quantum-mechanical analysis of band gaps and expected conductivity.
As a first step, we improved interatomic potentials for graphite and graphene to reproduce X-ray structure, density, cleavage energy, and contact angles according to experimental data, closing some gaps in existing models with deviations on the order of 30%. The parameters are embedded in CHARMM, PCFF, and other common force fields as part of the INTERFACE force field. The assembly of two peptides, selected by phage display, has been studied on graphene surfaces using molecular dynamics simulation in explicit water. The chosen peptides have the sequences EPLQLKM and HSSYWYAFNNKT. The thickness of graphene (1 to 5 layers) and the surface coverage with peptides (single molecules to multiple layers) have been varied to approximate experimental conditions in QCM. An analysis of binding residues, binding energies, conformations, and dynamic information of molecular mobility on the surfaces will be presented. Further analysis was carried out on the local scale by ab-initio calculations to understand trends in the band gap as a function of surface functionalization.

Deposition of cyclic azasilanes has been termed as “click chemistry on surfaces” due to the ease of reaction (<1min) and the lack of by-products during the deposition. Cyclic azasilanes are an advanced class of silane coupling agents which offer several advantages over the conventional silane coupling agents. Cyclic azasilanes are able to overcome the general limitations posed by conventional silanes which include need for catalytic amounts of water for the reaction to proceed, requirement of multiple hours to fully saturate a surface, and release of by-products upon reaction completion. The ring structure of cyclic azasilanes with Si and N centers adjacent to each other creates a strained structure which along with high oxophilicity of silicon contributes to the high reactivity of cyclic azasilanes towards available hydroxyl groups on any surface. As the size of the substrates shrinks into the nanometer regime, the surface energetics and the challenges involved in the accessibility of hydroxyl groups allows for the cyclic azasilanes to be superior in comparison to the conventional silane coupling agents. Thermodynamic and kinetic data will be reported for cyclic azasilanes surface modification.

The fabrication of organic coatings with well-defined structure and physico-chemical properties is of particular interest in several disciplines. Bio-organic layers on different length-scales, featuring variable composition, structure and modulus have been observed in several biological systems, such as human cartilage, mammalian skin and the nacre of oyster shells [1-3]. These complex materials consist of mechanically graded structures that resist and respond to the external normal and shear forces, protecting underlying tissues from incurring damage. Among the many “natural” coating systems, human epidermis is constituted by different layers of cells which present diverse properties to confer resistance against abrasion.
In order to mimic these natural structures, materials scientists have been made numerous efforts to fabricate coatings with graded and gradient-like mechanical properties within a single film. Despite this, the fabrication of a full-organic, polymer-based coating architecture featuring nano-scale variations of properties still represents a challenging task.
In order to fabricate polymer films presenting discontinuous and continuous variations of mechanical and chemico-physical properties we applied sequential surface-initiated polymerization (SIP) of different monomer compositions to create polymer brush/hydrogel films with vertically defined structure. Specifically, poly(hydroxyethyl methacrylate) (PHEMA) brush and brush-hydrogel layered films were synthesized by surface-initiated atom transfer polymerization (SI-ATRP) in the presence of different concentrations of diethylene and tetraethylene glycol dimethacrylate (DEG/TEGDMA). Sequential SI-ATRP of monomer mixtures was employed to fabricate brush-hydrogel films presenting vertically graded properties. Alternatively, continuous variation of monomers during the SI-ATRP resulted in brush-hydrogels featuring gradient-like variations of polymer architecture through the film thickness. The chemical, mechanical and tribological properties of the films were characterized by ellipsometry, FT-IR and colloidal probe atomic force microscopy (CP-AFM). All these measures confirmed the influence of polymer architecture on the properties of the films at specific depths.
Additionally, graded and gradient-like brush-hydrogels were used as reactors for the synthesis of polymer-inorganic hybrids presenting metal nanoparticles (NPs) embedded within the films. Depending on the vertical crosslinker content, different NPs morphologies were obtained at determined depths. Through this multi-step fabrication, polymer-inorganic hybrids with vertically defined structures and variable NPs loading were obtained. In conclusion we believe these methods will represent an easy and effective mean to form coatings with defined mechanical properties and tunable optical characteristics.
1. R. A. Stockwell et al., Nature 1967
2. J. C. Mackenzie, Nature 1969
3. H. D. Espinosa et al., Nat. Commun. 2011

Reentrant features on a surface are key to render it omniphobic, i.e., the apparent contact angle, theta;_r > 90°, especially when the intrinsic contact angle, theta;_o < 90°. However, a complete theoretical understanding of the wetting behavior of surfaces with reentrant features has remained unclear. Here, we present a unified and general model that can predict apparent contact angles for surfaces with reentrant features also. Unlike previous models, this model does not include the ‘roughness&’ of surfaces, but rather the specific geometry of the surface features (protrusions, cavities, etc.) to determine the apparent contact angle in terms of the
‘intrinsic&’ or ‘Young&’ contact angle (on a smooth planar surface) and two geometrydependent parameters. In particular, we find that cavities with reentrant walls (i.e., concave interior walls) result in important new conditions. The results show that both thermodynamic equilibrium and metastable states can arise, depending on the geometry (shape) and size of the cavities. We have fabricated some surfaces with micron-scale reentrant features (some of these structures mimic the surface texture of animals, birds, and plant leaves, e.g., lotus leaf, that render them hydrophobic or superhydrophobic), and have tested them with liquids of varying surface tensions, such as water, canola oil, and ethanol. We found that depending on the geometry of the surface features, the apparent contact angles span from the intrinsic angle to values that are very different and are in excellent quantitative agreement with the predictions of our model.

The control of gold nanoparticles&’ surface chemistry is a fundamental prerequisite to tailor their properties for biological applications such as bioimaging and sensing. Short peptides have been specifically designed to form self-assembled monolayers on gold nanoparticles surface^1,2 and used to increase their stability³ and enable functionalization with biomolecule at a stoichiometric level.^2,4 Here, we propose an experimental and computational approach to characterize the molecular organization and define the secondary structure of self-assembled monolayers of peptides on gold nanoparticles surface.
Experimentally, we are focusing on the benzophenone-derivative peptides, aiming to characterize the organization of mixed ligand shells on the gold nanoparticles surface. Under excitation at 350 nm, the carbonyl group of the benzophenone moiety crosslinks to an adjacent molecule. The cross-linking between the two adjacent ligands on the surface of the nanoparticles is monitored by Fourier Transform Infrared (FTIR), Ultraviolet-Visible (UV-Vis) spectroscopies and Mass Spectrometry (MS), providing insight into the molecular organization and structural response of these ligand shells upon light irradiation.
We are using Molecular Dynamics simulations to investigate the secondary structure of two short peptides, i.e. CALNN and CFGAILSS, on spherical GNPs of different sizes, i.e. 5, 10, 25 nm. The model is validated on the basis of Shaw et al. work⁵ where they have shown, using a combination of Fourier Transform Infrared (FTIR) spectroscopy and Solid-State Nuclear Magnetic Resonance (SSNMR), that the CFGAILSS&’ conformation on the gold nanoparticles surface depends on the size of the particle, hence on its curvature.
Both these approaches are necessary in order to have an insight at the molecular level into the organization and structure of self-assembled monolayers of peptides coating the gold nanoparticles surface.
References:
1. Lévy, R. et al. Rational and combinatorial design of peptide capping ligands for gold nanoparticles. J. Am. Chem. Soc.126, 10076-84 (2004).
2. Duchesne, L., Gentili, D., Comes-Franchini, M. & Fernig, D. G. Robust Ligand Shells for Biological Applications of Gold Nanoparticles. Langmuir24, 13572-13580 (2008).
3. Chen, X. Y. et al. Features of Thiolated Ligands Promoting Resistance to Ligand Exchange in Self-Assembled Monolayers on Gold Nanoparticles. Aust. J. Chem.65, 266-274 (2012).
4. Duchesne, L. et al. Transport of fibroblast growth factor 2 in the pericellular matrix is controlled by the spatial distribution of its binding sites in heparan sulfate. PLoS Biol.10, 16 (2012).
5. Shaw, C. P., Middleton, D. a, Volk, M. & Lévy, R. Amyloid-derived peptide forms self-assembled monolayers on gold nanoparticle with a curvature-dependent β-sheet structure. ACS Nano6, 1416-26 (2012)

The oriented migration of cells is fundamental to many physiopathological processes, including in muscle regeneration. Cellular motion is guided by extracellular gradients of signaling proteins, so-called chemokines. A family of linear polysaccharides, known as glycosaminoglycans (GAGs) helps in organizing and maintaining the haptotactic gradients of chemokines on the cell surface and in the extracellular matrix, thus providing directional cues for migrating cells.
The ability to control and characterize the supramolecular presentation of GAG chains and their interaction with chemokines is to date very limited, especially when combined with ECM components. Our approach consists in designing biomimetic surfaces, reconstituting heparan sulfate (HS, an example of GAG), and other cell membrane and ECM components (e.g. the peptide sequence RGD (Arg-Gly-Asp) as a cell adhesion ligand), into tailor-made and multifunctional model surfaces. Despite strong indications for their functional importance, HS has so far been largely neglected in in vitro models due to its limited availability in sufficiently pure and suitably functionalized form, and a lack of methodologies to integrate it into assemblies. Here, we present the functionalization of HS with biotin at the reducing end via oxime ligation. This facile, one-step, high yield and broadly applicable method leads to highly stable terminally functionalized GAGs, which is superior compared to the most frequently used hydrazone ligation strategy. RGD was also biotinylated and grafted along with HS onto streptavidin-modified oligo ethyleneglycol (OEG) monolayers and supported lipid bilayers (SLBs) that constitute our model surfaces. Quartz crystal microbalance (QCM-D) and ellipsometry permitted us to characterize and control the supramolecular presentation of HS and RGD - the local density, orientation and conformation - and their interaction with a selected chemokine, stromal derived factor 1 (SDF-1α/CXCL12). The kinetics of chemokine binding to HS was quantified using surface plasmon resonance (SPR). These well-defined model surfaces were used to study the response of myoblasts to chemokines presented via HS. Myoblasts were found to respond to HS-bound SDF-1α through enhanced adhesion, spreading and motility, on surfaces presenting only HS-bound SDF-1α but also on surfaces co-presenting RGD.
In conclusion, novel tailor-made biomimetic surfaces were engineered, displaying several types of biomolecules at controlled orientation and mean surface density. These novel biomimetic surfaces could potentially generate various insights in the field of glycomimetics, e.g. in unravelling the function of HS in the chemokine-mediated migration of myoblasts. These model surfaces also permit a controlled presentation of SDF-1α which holds a key importance in the design of SDF-1α-loaded implantable devices, for regenerative medicine and tissue remodeling strategies.

Interfaces between synthetic materials and biological systems (bio- interfaces) constitute one of the most dynamic and expanding fields in biomaterials science. It is virtually known that upon contact of cells or tissues with biomaterials, selective and competitive absorption of key serum proteins on bimaterial surfaces is the initial event participating in cell-biomaterial interactions. Conformational changes associated with the adsorption of the proteins play crucial roles in determining the dynamic behaviors of cells, such as adhesion, proliferation, differentiation, etc, on biomaterial surfaces. Structural information of the absorbed proteins is therefore essentially required for a comprehensive understanding at molecular level of how cell-biomaterial interactions function. This paper presents our recent research progress in visualizing the conformational changes of several important serum proteins, i.e. fibronectin, album, vitronectin, after their adsorption on the surfaces of nanostructured hydroxyapatite/graphene and diamond-like carbon films. Single particle electron microscopy and atomic force microscopy were both employed for the investigation. Preferred adsorption of the proteins like fibronetin on graphene etc has been revealed. And varied conformations of the proteins were disclosed depending on topographical morphology and surface chemistry of the material samples. The conformations acquired from the EM/AFM observations presumably account for the state of the spreading and subsequent proliferation of the cells on the surfaces of the typical biomedical nanocomposites. The results shed light on understanding the bio-interfacial science that involves manufacturing and characterization of functional surfaces for specific interactions with bio-systems and the molecular and kinetic processes occurring at such interfaces.

The interactions of biomolecules with human tissue structures provide a rich #64257;eld of inquiry, with fundamental importance to a variety of biological and medical sciences. For instance, the ability to synthesize or isolate target biomolecules allows us to engineer certain desirable e#64256;ects and reactions. This has resulted in the inclusion of a large family of biomolecular components in a multitude of commercial products and medical treatments. However, the full complexity of these biomolecular interactions is not always fully understood; this lack of knowledge can result in everything from sub-optimum formulations to (in the worst case) harmful unintended side e#64256;ects.
In this work, we consider the interactions of one common class of commercially and medically relevant biomolecules, namely antioxidants. Speci#64257;cally, we select the well-known biomolecular antioxidant species α-tocopherol acting on the components of human stratum corneum for study. We choose stratum corneum because 1) this represents a practically relevant scenario and 2) stratum corneum has a well-studied lipid-protein composite structure. We examine the impacts of topical application of such an antioxidant on these components. Using mechanical thin #64257;lm techniques including double-cantilever beam fracture tests (DCB) and substrate curvature experiments as well as attenuated total re#64258;ectance Fourier transform infrared spectroscopy (ATR-FTIR), we measure the di#64256;usion rate, e#64256;ect on cohesion energy, and UV protective impacts of α-tocopherol on human tissue. Analysis of these data suggests that lipophilic biomolecular antioxidants such as tocopherol partake in lipid interactions which serve to modify the biomechanical behavior of skin tissue, for instance reducing the critical intercellular cohesion energy. These results have implications for the future design of improved topical treatments, as well as fundamental impact on our understanding of this class of biomolecules and their interactions.