The synergistic combination of nanotechnology and biological has resulted in numerous innovative approaches for therapy and biology. One of the most exciting prospects of nanotechnology is that nanoparticles can act as a handle by which one can control biological processes. Unfortunately, one of the biggest barriers for the effective use of nanoparticles in biology is non-specific adsorption, where proteins and DNA non-covalently stick to nanoparticles. As soon as nanoparticles are introduced to a biological fluid, the proteins and other species that are present at high concentration non-specifically adsorb to their surfaces, forming a “protein corona.” This corona is impossible to prevent and can block the surface of the nanoparticle, as well as cause undesired side effects in targeting and biodistribution. Furthermore, non-specific adsorption is complex and difficult to control, it is typically viewed as a major hindrance to nanobiotechnology. Despite this, protein coronas have unique characteristics that can impart desirable properties to nanoparticles. We demonstrate protein adsorption to gold nanoparticles can enhance in vitro translation. In addition the protein corona can be used in triggered release. We form protein coronas around gold nanorods such that they can hold large quantities of small molecules and DNA at a capacity higher than what is achievable by covalent attachment strategies. The nanorods can be excited by laser irradiation on their SPR, which heats the nanorods and disrupts the corona, releasing the drug or DNA. We are using triggered release of species from the gold nanorods for a variety of biological and therapeutic applications.

Vertically aligned arrays of silicon nanowires (SiNWs) provide a multifunctional platform for controlling stem cell differentiation with nanoscale topography. It has been shown that the spacing between the NWs can influence stem cell morphology [1], and by using NWs in an array to penetrate the cell membrane, biomolecules can be delivered into the interior of a cell [2]. In addition to the nanoscale topography, surface chemistry plays a critical role in determining cell fate. In this work, we use metal-assisted chemical etching to produce porous SiNWs with different surface treatments for controlling mesenchymal stem cell growth. SiNWs were fabricated with a variety of diameters, pitch, and surface treatments including native oxide, poly-D-lysine and laminin, and atomic layer deposition (ALD) of Al₂O₃. The growth of mesenchymal stem cells was characterized up to 2 weeks after cell plating. While UV-ozone treated SiNWs with a native oxide showed almost no mesenchymal stem cell survival, SiNWs coated with ALD Al₂O₃ yielded survival rates similar to the control substrates. These results will be discussed in light of our goal to understand the role of SiNW geometry and surface chemistry in determining stem cell growth.
[1] M. Bucaro, Y. Vasquez, B.D. Hatton, J. Aizenberg, ACS Nano6, 6222 (2012).
[2] A. Shalek et al. Proc. Natl. Acad. Sci. USA107, 1870 (2010).

Our objective is to design nanostructured hybrid inorganic-biological
materials using the self-assembly of functionalized nanotubes and
lipid molecules. In this presentation, we summarize the multiple
control parameters which direct the equilibrium morphology of a
specific class of nanostructured biomaterials. Individual lipid
molecules are composed of a hydrophilic head group and two hydrophobic
tails. A bare nanotube encompasses an ABA architecture, with a
hydrophobic shaft (B) and two hydrophilic ends (A). We introduce
hydrophilic hairs at one end of the tube to enable selective transport
through the channel. The dimensions of the nanotube are set to
minimize its hydrophobic mismatch with the lipid bilayer. We use a
Molecular Dynamics-based mesoscopic simulation technique called
Dissipative Particle Dynamics which simultaneously resolves the
structure and dynamics of the nanoscopic building blocks and the
hybrid aggregate. The amphiphilic lipids and functionalized nanotubes
self-assemble into a stable hybrid vesicle or a bicelle in the
presence of a hydrophilic solvent. We demonstrate that the morphology
of the hybrid structures is directed by factors such as the
temperature, the molecular rigidity of the lipid molecules, and the
concentration of the nanotubes. We present material characterization
of the equilibrium morphology of the various hybrid nanostructures. A
combination of the material characterization and the morphologies of
the hybrid aggregates can be used to predict the structure and
properties of other hybrid materials.

DNA origami is a relatively new technique that uses hundreds of short “staple” DNA oligonucleotides to direct the folding of a single stranded DNA scaffold into a predefined structure. Such folded DNA structures possess nanometer precision and can be used as high precision templates for nanofabrication. Here we present the use of DNA origami as a template for arranging nano objects into helical configuration. Unlike previous works that uses single DNA origami as template, we assemble multiple DNA origami monomers into larger nano-helices whose dimensions approach the wavelength of visible light. Such larger structures are capable to template more nano objects with larger dimensions and can lead to stronger chiral properties. We also demonstrate that the chirality of the nano-helices can be readily controlled by rationally design the complementary DNA strands that link adjacent DNA origami monomers together. The strategy we presented here offers a facile way to create nano helices and tune their unique properties of these chiral structures.

Numerous artificial transport systems utilizing carriers, channel-forming or self-organized polymeric superstructures able to orient, to select and to pump the ionic transport across membranes have been developed in the last decades. Of special interest is the structure-directed function of hybrid membrane materials and control of their build-up from suitable units by self-organisation. The main interest focus on functional hybrid membrane materials in which the recognition-driven transport properties or the creation of directional transporting pathways could be ensured by a well-defined incorporation of receptors of specific molecular recognition and self-organization functions. We are therefore proposing to review the membrane transport properties of such supramolecular hybrid materials with potential in renewable energy applications as fuell-cells technology, biomimetic water and ion channels, etc.
[1] M. Barboiu, C. Luca, C. Guizard, N. Hovnanian, L. Cot, G. Popescu, J. Membrane Sci., 1997, 129, 197-207.
[2] M. Barboiu, C. Guizard, J. Palmeri, C. Reibel, C. Luca, L. Cot, J. Membrane Sci. 2000, 172, 91-103.
[3] M. Barboiu, S. Cerneaux, G. Vaughan, A. van der Lee, J. Am. Chem. Soc. 2004, 126 3545-3550.
[4] A. Cazacu, C. Tong, A. van der Lee, T.M. Fyles, M. Barboiu, J. Am. Chem. Soc. 2006, 128(29), 9541-9548.
[5] C. Arnal-Herault, A. Pasc-Banu, M. Barboiu A. van der Lee, Angew. Chem. Int. Ed. 2007, 46, 4268-4272.
[6] C. Arnal-Herault, A. Pasc-Banu, M. Michau, M. Barboiu, Angew. Chem. Int. Ed. 2007, 46, 8409-8413.
[7] C. Arnal-Hérault, M. Barboiu, A. Pasc, M. Michau, P. Perriat, A. van der Lee, Chem. Eur. J. 2007, 13, 6792
[8] M. Michau, M. Barboiu, R. Caraballo, C. Arnal-Hérault, A. van der Lee, Chem. Eur. J. 2008, 14, 1776-1783.
[9] M. Barboiu, P. Aimar, J.M. Lehn, From simple molecules to complex membrane systems, Editorial, Special issue of J. Memb. Sci., 2008, 321, 1-2.
[10] M. Michau, M. Barboiu, J. Mater. Chem., 2009, 19, 6124-6131.
[11] A. Cazacu, Y.M. Legrand, A. Pasc, G. Nasr, A. van der Lee, E. Mahon, M. Barboiu, Proc. Natl. Acad. Sci., 2009, 106(20), 8117-8122.
[12] S. Mihai, A. Cazacu, C.Arnal-Herault, G. Nasr, A. Meffre, A. van der Lee, M. Barboiu, New. J. Chem, 2009, 33, 2335-2343.
[13] M. Barboiu, Chem. Commun. 2010, 46, 7466-7476.
[14] Y. Le Duc, M. Michau, A. Gilles, Valerie Gence, Y.-M. Legrand, A. van der Lee, S. Tingry, M. Barboiu,; Angew. Chem. Int. Ed. 2011, DOI : 10.1002.anie.201103312

Genetically engineered protein-based polymers (GEPBP) expand the sophisticated multifunctional properties of biopolymers with the precise incorporation of functionalities. Elastin-like polypeptides (ELPs) populate the lexicon of GEPBPs by virtue of their unique stimuli-responsive, mechanical, and bio-compatible properties. A central concept of our hybrid inorganic-biomaterial is to construct a library of metal-biopolymer complexes by combining the rich information content of ELPs with the tunable optical, redox, magnetic, and chemical properties of metal-ligand units. From this strategy, we have designed and prepared three related classes of transition-metal complexes with polypyridyl derivatives. In class I, we have adopted a “plug and play” approach where the side-chain length of functional groups has been varied systematically to procure a spectrum of reactive ligands with variable exposure. In class II, we have used alterations in substituent groups to modulate the strong metal-to-ligand charge-transfer absorptions and associated redox potentials of the complexes. In class III, we have developed a suite of complexes in which one fixed ligand has conjugation sites while the others are varied to achieve improved photophysical and redox properties that can be tunable by conditions of the medium, such as pH or solvent. We have implemented carbodiimide-based coupling chemistries to take advantage of our new ligands functionalized with carboxyl or amino groups complementary to the side-chain functionalities of ELP units. The successful introduction of metal-ligand components into specific protein sites has been confirmed by several techniques, including gel electrophoresis and time-dependent monitoring of the bioconjugation reactions using mass spectrometries. We have also been able to characterize these bio-conjugates in a library of ELPs to evaluate their functional properties. We anticipate that this approach to (bio)polymeric architectures will lead to advanced stimuli-responsive materials for a broad range of applications.

Hybrid inorganic-biological materials have great potential but also unique challenges, including biocompatibility, functional integration at multiple length scales, and responsiveness to external stimuli. Human skin can form a particularly interesting inorganic-biological construct when exposed to selected treatments involving active molecules or second phase inorganic particles. For example, to protect against the harmful effects solar ultraviolet (UV) exposure, inorganic UV-blocking micron- and nano-sized zinc oxide (ZnO) and titanium dioxide (TiO2) particles are commonly applied to the outermost layer of skin, the stratum corneum (SC). These diffuse and partition into the SC intercellular boundaries, and the resulting inorganic-biological construct is highly effective in preventing damage and disease from solar UV radiation. However, it remains unclear precisely how these treatments interface with the biological components and if these treatments can prevent alterations to the biomechanical barrier of the SC.
We explored the interaction of zinc oxide and titanium dioxide particles with SC in the presence of UV radiation. We used substrate curvature techniques to characterize the drying stresses, and hence the driving force for damage, that occur with UV exposure and the ability of inorganic particles to mitigate this damage. We also explored the ability of the particles to protect the innate SC resistance to corneocyte separation, which has been shown to decrease under UV exposure. We found that the inorganic UV inhibitors protected the SC&’s mechanical properties under relatively large doses of UV radiation. We also compared efficacy of the inorganic molecules to chemical (UV absorbing) sunscreens. Clinical implications of this work include prevention and treatment of sunburn and long term skin damage such as photoaging.

The application of nanoplasmonic materials relies on the fabrication of metallic nanostructures with well defined sizes and geometries. For this purpose, various chemical approaches have been developed to synthesize metal nanoparticles, in particular gold nanoparticles (AuNPs). [1] For many applications the plasmon resonance wavelength needs to be adjusted to the wavelength of the exciting light source. This can be achieved by tailoring the particle morphology or by assembling pre-synthesized AuNPs into networks, whose plasmon resonance is governed by interparticulate coupling. Via DNA-hybridization, extended networks can be formed that can be tailored by the interconnecting DNA and the ratio of reactants. [2] These systems can be reversibly decomposed photothermally via temperature-induced DNA-dehybridization, with possible applications in fields such as drug release or gene regulation. [3]
However, many applications of plasmonic materials, e.g. in imaging techniques, require finite size structures. [4] In contrast to extended networks that exhibit long range orders in space, thus yielding iso-structural X-Ray diffraction patterns, we studied spatially defined networks using dynamic light scattering, small angle X-ray scattering (SAXS) as well as optical extinction spectroscopy. [5] In this presentation we will show for the first time a combination of these techniques with in situ-TEM and Mie simulations to analyze the internal structure of finite sized assemblies. Different interparticle distances are identified and quantified within these assemblies. By controlling the network morphology, centre-shell geometries are obtained and a model to determine, from the SAXS data, the number of shell AuNPs is developed.
The correlation of optical characteristics with structural features enables the quantitative description of such DNA-AuNP hybrid systems. This description is expected to prove useful for the construction and application of such systems for example in drug release, gene regulation or external stimuli responsive materials.
[1] Timper et al., Angew. Chem. 2012, 124, 7705; Goesmann et al., Angew. Chem. 2010, 122, 1402.
[2] Mirkin et al., Nature 1996, 382, 607; Alivisatos et al., Nature 1996, 382, 609; Park et al., Nature 2008, 451, 553; Maye et al., Small 2007, 3, 1678.
[3] Rosi et al., Science 2006, 312, 1027; Braun et al., ACS Nano 2009, 3, 2007.
[4] Gutrath et al., Nanotechnology 2012, 23, 225707.
[5] Buchkremer et al., Small 2011, 7, 1397.

The widespread use of silver nanoparticles (Ag-NPs) in consumer and medical products provides strong motivation for a careful assessment of their environmental and human health risks. Recent studies have shown that Ag-NPs released to the natural environment undergo profound chemical transformations that can affect silver bioavailability, toxicity, and risk. Less is known about Ag-NP chemical transformations in biological systems, though the medical literature clearly reports that chronic silver ingestion produces argyrial deposits consisting of silver-, sulfur-, and selenium-containing particulate phases. Here we show that Ag-NPs undergo a rich set of biochemical transformations, including accelerated oxidative dissolution in gastric acid, thiol binding and exchange, photoreduction of thiol- or protein-bound silver to secondary zerovalent Ag-NPs, and rapid reactions between silver surfaces and reduced selenium species. Selenide is also observed to rapidly exchange with sulfide in preformed Ag2S solid phases. The combined results allow us to propose a conceptual model for Ag-NP transformation pathways in the human body. In this model, argyrial silver deposits are not translocated engineered Ag-NPs, but rather secondary particles formed by partial dissolution in the GI tract followed by ion uptake, systemic circulation as organo-Ag complexes, and immobilization as zerovalent Ag-NPs by photoreduction in light-affected skin regions. The secondary Ag-NPs then undergo detoxifying transformations into sulfides and further into selenides or Se/S mixed phases through exchange reactions. The formation of secondary particles in biological environments implies that Ag-NPs are not only a product of industrial nanotechnology but also have long been present in the human body following exposure to more traditional chemical forms of silver.

Magnetic nanoparticles (MNPs) are essential to many technologies, from the storage of electronic data, to their use in biomedical applications to treat cancer. It is essential that the magnetic behavior of these MNPs is reliably consistent, which means that they must have a monodispersed size and shape distribution. To synthesise uniform MNPs usually requires heating toxic reagents to high temperatures, which is not very environmentally friendly. Genetically encoded biomineralization proteins are able to control the formation of about 60 different biomaterials in nature [1]. They are able template specific crystallographic phases to form nanoparticles and complex inorganic architectures, all under physiological conditions. Some of these biomineralization proteins, as well as smaller biotemplating peptides, are also able to direct aqueous reagents to form consistent, uniform nanoparticles under mild reaction conditions in vitro [2]. Thus, the use of biotemplating offers a far greener approach to the synthesis of nanoparticles for use in applications.
To be used for data storage, MNPs must be attached onto a surface, often in patterns or even as a thin-film. This usually involves the use of clean-rooms, photolithography and sputtering under vacuum, which is costly and extremely energy intensive. Soft-lithographic techniques, such as micro-contact printing, have been used to functionalize patterned surfaces with biomineralizing proteins and peptides. These patterns have then been used to biotemplate the formation of thin-films of materials and nanoparticles. In recent work we showed that the biomineralizing protein Mms6 is able to template the formation of high quality magnetite nanoparticles when selectively immobilised onto surface [3]. This was the first time that magnetic materials have been biotemplated onto surfaces. As magnetite is magnetically soft (i.e. has a low coercivity), it is unlikely that the Mms6 templated magnetite MNPs could be used for data storage, as any information written onto a soft material is likely to be easily lost. As such, methods to create biotemplated MNPs on patterned surfaces with a much higher coercivity will be discussed. By using these bioinspired approaches, we should be able to create arrays of biotemplated magnetic materials under mild reaction conditions that may be suitable for use in data storage applications.
[1] RA Metzler et al. (2008) Langmuir, 24, 2680-2687.
[2] JM Galloway & SS Staniland (2012) J. Mater. Chem., 22, 12423-12434.
[3] JM Galloway et al. (2012), Small, 8, 204-208.

Down-sized inorganic materials have unique functions, but the manipulation of pattering and assembling them still remains difficult. Recently, artificial peptides with affinity for nonbiological inorganic materials have been identified, and we generated the antibodies with high affinity for inorganic material surfaces by the combination of peptide grafting and in vitro molecular evolution methods. The material-binding biomolecules are expected to be used for bottom-up fabrications of nano-materials. In this study, we utilized the material-binding antibodies to functionalize nano-materials, and bispecific anti-material antibodies were constructed as bio-interface molecules to assemble nanomaterials.
To apply the high affinity of material-binding antibody fragments to stabilization of nano-particle, the antibodies with affinity for zinc oxide (ZnO) (4F2 VHH) were added to a ZnO particle suspension, respectively. In general, ZnO particles used are immediately precipitated; however, 4F2 VHH functioned as dispersant in comparison with bovine serum albumin (BSA). When we mixed gold-binding antibodies (AuE32 VHH) and gold nanoparticles, the particles were dispersed even in a solution with 1 M NaCl, where gold nanoparticle with BSA were immediately aggregated. These results indicate that inorganic nano particles were highly stabilized by the binding of material-binding antibodies on material surfaces.
In addition, we attempted to create the artificial antibody with bispecificity for ZnO and Au materials by connecting 4F2 VHH and AuE32 VHH via a hinge-linker to propose the application of material-binding antibodies as bio-interface molecules. When the bispecific antibodies were added to the mixture solution of ZnO and gold particles, the plasmon absorbance derived from gold particles were drastically decreased. The linkages between ZnO and gold particles via the bispecific antibodies induce the aggregation of gold particles so that the plasmon absorbance was decreased. Thus, we succeeded in showing the potential of anti-material antibodies for manipulating nano-materials.

We developed a simple and robust method to pattern the attachment of biomolecules to a planar inorganic surface. The technique is based on imprinting the surface with a self-assembled colloidal crystal. The result of the imprinting is a triangular array of reactive sites with sub-micron periodicity that covers square centimeters of surface area. Biomolecules can then be covalently immobilized on the imprinted array through standard amine - carboxylic acid conjugation methods.

The discovery of the principle molecules involved in biosilification both in diatoms (silaffins and polyamines) and sponges (silicateins) brings out a new paradigm for silica synthesis at environmentally benign conditions. Silicatein has high sequence identity and similarity with that of cathepsin L in which the cysteine at the active site is replaced by serine in silicatein. Previous reports showed human cathepsin L was able to condense silica by changing the cysteine into serine residue at its active site. In this study, we obtained hypothetical cathepsin-like protein (CAT) from Nematostella vectensis which displays 55 % identity and 75% similarity with mature silicatein alpha (SIL) of Suberites domuncula from a BLAST-based search of protein sequence. This protein expressed in E.coli displayed not only N-carbobenzoxy-L-phenylalanyl-L-arginine 4-methylcoumaryl-7-amide (Z-FR-AMC) hydrolytic activity but also silica condensing activity. To increase the silica forming activity, some residues including cysteine in active site were changed into silicatein conserved residues, resulting in 65 % identity and 79 % similarity with SIL. The mutant silicatein-like cathepsin (SLC) revealed increased expression level in E. coli and silica forming activity which was comparable to that of SIL and decreased protease activity as compared to those of CAT. Whereas the expression of CAT in E. coli was not stable, that of SLC was stable. The solubility of SLC was increased compared to that of SIL. SLC produced silica particles of size less than 50 nm which were increased to 200~300 nm in the presence of a structure-directing agent, Triton X-100. Bovine carbonic anhydrase was efficiently immobilized by simple adding it to the reaction mixture during SLC-mediated silification with tetraethyl orthosilicate as a substrate at near-neutral pH in an aqueous solution at ambient temperature. Immobilized protein retained its enzymatic activity for longer time and was reused up to several times. In conclusion, hypothetical cathepsin-like protein from Nematostella vectensis was evolved to more soluble and available biosilica forming protein and SLC-mediated biosilica can be applied for various silica-based materials.

A novel optical whole cell-based array biosensor for detection of neurotoxic paraoxon was developed by immobilizing recombinant Escherichia coli expressing organophosphorus hydrolase in periplasmic space on mussel adhesive protein (MAP)-coated 96-well microplate. The effect of MAP as a cell immobilizing agent was evaluated by comparing with simple physical adsorption-based cell immobilization. Paraoxon-degradable activity and fluorescence microscopy studies revealed that the use of MAP has advantages of increasing cell immobilizing efficiency and enhancing stability of immobilized cells compared to conventional simple physical adsorption-based whole cell biosensor. Scanning electron microscopy analysis also showed that MAP effectively immobilized cells on surface without pretreatment steps. The whole cell-based array biosensor prepared by using optimal quantities of the cell loading (4 OD600) and MAP (50 mu;g/cm2) could detect as low as 5 mu;M paraoxon with high reproducibility (relative standard deviation is 3.92% for n = 19) and the detection range of the biosensor was 5-320 mu;M. In addition, the whole cell biosensor showed good long-term storage for one month with the retention of 80% activity and multiple-use stability. Collectively, our proposed novel whole cell array biosensor is suitable tool for rapid and sensitive detection of harmful organophosphorus compounds.

In recent years, the availability of low cost, renewable energy sources capable of meeting a significant portion of global energy demands has become a concern. Unlike many second generation solar cell materials which contain scarce and possibly toxic elements, Cu₂S is an abundant, non-toxic material. Furthermore, its 1.2 eV bandgap is near the theoretical optimum for maximum efficiency of a single junction solar cell. Historically, Cu₂S-based cells have been problematic due to rapid Cu diffusion caused by elevated fabrication temperatures; however our biomineralization approach occurs at room temperature thereby minimizing diffusion. In this work, we investigate the optical and electrical behavior of nanocrystalline Cu_xS nanowires biomineralized with a M13 viral template. The filamentous geometry of the M13 virus, approximately 880 nm in length and 6 nm in diameter, is particularly well-suited for nanowire synthesis and electrical characterization. To enhance the affinity of the template for Cu_xS mineralization, the M13 phage was genetically modified to display a peptide fusion of three glutamic acid residues at the N-terminus of each of 2700 copies of the p8 protein found along its length. The addition of acidic amino acids to the M13 major coat protein increased the net negative charge of the template such that when it was incubated with CuCl₂ large bundles of phage developed rapidly. The formation of these fiber-like bundles is consistent with previous reports by other researchers and was likely caused by strong non-specific, electrostatic interactions between the highly negatively charged genetically-modified template and the positively charged divalent Cu ions within the CuCl₂ precursor solution. The phage/copper ion bundles were separated through centrifugation, the CuCl₂ removed, and Na₂S added as a S source to convert the Cu ions bound along the length of the virus to Cu_xS. The mineralized virus bundles appeared as brown fiber-like structures in solution. Very large agglomerates of phage, as well as smaller clusters composed of only a few phage and individual mineralized phage were observed with TEM. Nanocrystals were located along the length of the viral-template regardless of the bundle size. EDX spectroscopy was used to verify the presence of Cu and S within the inorganic material. The distinct rings observed in the electron diffraction pattern confirmed the crystallinity of the biomineralized material and the calculated d-spacings were attributed to crystal planes within the cubic Cu₂S and Cu_1.8S structures. The effective bandgap of the biomineralized nanocrystalline Cu_xS was determined from the UV/VIS/IR absorption spectrum. The longitudinal electrical performance of the nanowires will be characterized using two-terminal measurements. Furthermore, the photoelectrical response will be studied with wavelength specific photocurrent action measurements, as well as broadband illumination measurements.

Biocompatible core-shell structured hybrid nanoparticles (NPs), which combine optical properties of a metal nanoshell with the size, shape and attractive properties of a nano-sized polymer core, have emerged as promising nanodevices for high-sensitivity cancer detection and treatment. Among metallic NPs, the bimetallic Au-Ag NP is most attractive due to its high absorption capability and plasmon resonances in the UV-vis-NIR range, which makes it uniquely suitable for many biomedical applications. NPs based on surface enhanced Raman scattering (SERS) are a new class of tags for biodetection including cancer detection. For Au-Ag nanostructures, a signal intensity enhancement factor of 104-106 may be achieved as a result of the signal increase in gaps between the center and sharp tips of a flower-like Au-Ag NP, which is sufficient for obtaining Raman signals large enough to enable single molecule detection. In the present study, two strategies, using either an Au-Ag nanoshell or a nano-sized Au-Ag core, were investigated to create two types of nanodevices for potential cancer detection and treatment applications. The first type was core-shell structured, anti-cancer drug-loaded poly(lactide-co-glycolide) (PLGA)@Ag-Au NPs. Monodispersed, Paclitaxel (PTX)-loaded PLGA NPs were made using a stabilizer-free method, and the PTX encapsulation efficiency approached 95%. A porous Ag nanoshell was then formed in situ on PTX-loaded PLGA NPs through the controlled reduction of AgNO3 by polyvinylpyrrolidone. The NPs thus formed were used to grow Ag-Au nanoshells on the PLGA core in a replacement reaction. In this nanodevice, the bimetallic Ag-Au nanoshell provided SERS enhancement, and in vitro SERS experiments using 4-MBA as the Raman reporter showed that the PLGA@Ag-Au hybrid NPs greatly amplified Raman signals, rendering them as highly desirable SERS optical tags for cancer detection. The antibody anti-HER2 could be conjugated to the nanodevice, enabling it to target HER2-overexpressing SK-BR-3 cancer cells. The hyperthermia effect caused by the Au-Ag shell under NIR-irradiation and drug released from the PLGA core would provide cancer treatment. The second type used highly branched Au-Ag NPs as the core and coated them with a layer of folic acid-chitosan (CS-FA) conjugates. Compared to simple Au or Ag NPs, bimetallic Au-Ag NPs displayed higher SERS. The receptor for FA constituted a useful target for tumor specific detection, which would also facilitate internalization of NPs through cancer cell membrane. Therefore, the nanodevices created were able to target folate receptor over-expressing Hela cells through the folic acid on the nanodevice surface. The hyperthermia effect caused by the Au-Ag core under NIR-irradiation would provide cancer treatment.

The continuous spread of microbial pathogens with antibiotic-resistance has led to a growing interest in the design and development of new materials that are effective in elimination of bio-hazards. Photodynamic Inactivation (PDI) is a promising approach for elimination of antibiotic-resistant bio-hazards. Targeting of the photosensitizer (PS) to the bio-hazards is required to improve efficiency and avoid possible damage of host tissues. In this study, we synthesized the multifunctional nanoparticle (Fe3O4@PS/AP) conjugated with photosensitizers and acceptor proteins (AP) and characterized to check the application possibility as a new and effective antimicrobial reagent. The multifunctional nanoparticles are capable of selective elimination of bio-hazards from aqueous solution. As the specific bio-hazards are exposed to the appropriate wavelength (510 nm), the multifunctional nanoparticles localized to the bio-hazards surface effectively eliminate them at a low power density light. Such novel multifunctional nanoparticle can be applied in various medical applications, such as in magnetic resonance imaging (MRI) diagnosis, bio-separation, target delivery, and PDT.

Presence of nanoparticles such as gold and silver in high concentrations in environmental is causing adverse health issues due to their high toxicity1-5. Therefore, it is important to develop new water treatment methods to remove nanoparticles. Use of low cost biomembranes for removal of nanomaterials is interesting due to high accessibility and low cost. Biomembranes loaded with zirconium (IV) oxide have been used to extract PVP capped gold and silver from aquesous environment. Immobilization of zirconium onto the membrane surface facilitated the extraction of nanoparticles. Structural modifications of the adsorbent were characterized using SEM, EDS and XPs. Removal efficiency was estimated using batch adsorption studies. Adsorption kinetics for all pollutants at different concentrations are described in terms of pseudo-second-order rate equation with respect to adsorption capacity and correlation coefficients.
Acknowledgement: The authors thank the Environment and Water Industry Programme Office (EWI) under the National Research Foundation of Singapore (PUBPP 21100/36/2, NUS WBS no. R-706-002-013-290, R-143-000-458-750, R-143-000-458-731) for the financial support of the work. RM also thanks the National University of Singapore for a scholarship for graduate studies.
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This talk will demonstrate a novel fabrication process for enclosed graphene chambers to encase liquids, living cells and solution-dispersed nanomaterials. These encasements will enable liquid phase imaging via electron-microscopes. Here, the strongly-repelling Π clouds in the interstitial sites of graphene&’s lattice makes it impermeable, graphene&’s high electron-transparency makes for unsuppressed resolution, and the C-C bond flexibility enables conformal encasement. Moreover, graphene&’s high Young&’s modulus retains the structural integrity under TEM conditions, while its high electrical and thermal conductivity significantly abates electrostatic-charging. For example, bacterial cells can be encased within graphenic chamber to preserve their dimensional and topological characteristics under high vacuum (10-5 Torr) and beam current (150 A/cm2). We will also demonstrate solution-dispersed nanoparticles within micron-scale graphene chambers. We will also demonstrate the selective creation of wrinkles on graphene using bacteria as a scaffold.

We describe here the growth of cadmium sulfide (CdS) nanotubes, templated and assembled on microtubule (MT) templates. The controlled growth and organization of CdS nanostructures is critical if the valuable optical, electronic, and chemical properties of this wide band gap semiconductor are to be fully utilized in emerging technologies. Polymerized from tubulin dimers, MTs are tubular proteinaceous nanofilaments only 25 nm in diameter, but extending in length to over a millimeter, and they offer unique opportunities to template CdS nanostructures. In cells, these chemically and functionally rich biological nanofibers interact with other intracellular machinery to facilitate and direct the complex organization of nanomaterials. Here, we describe a bio-inspired, bio-mediated approach to CdS nanotube growth, exploiting these natural organizing elements as templates. We utilize a biomimetic synthesis to grow uniform cubic zincblende CdS nanocrystals on MTs, replicating the MTs&’ tubular morphology with dense CdS only a single nanocrystal thick. We additionally exploit specific interactions of MTs with microtubule associated proteins to direct the organization of these MT templates, enabling the subsequent directed growth of semiconducting CdS nanotubes into specific bio-mediated architectures including linear arrays, three-dimensional asters, and rings. These MT-mediated demonstrations of nanoscale materials synthesis and assembly open the door to a promising new level of complexity and control over materials templating that may be achieved using such biological tools and processes.
Sandia National Laboratories is a multi program laboratory managed and operated by Sandia Corporation, a wholly owned subsidiary of Lockheed Martin Corporation, for the U.S. Department of Energy's National Nuclear Security Administration under contract DE-AC04-94AL85000.

Storage and transport of biological samples for long-term studies costs millions of dollars per year. Silica encapsulation (non-cryogenic) is a promising method for long-term storage and transport of biological materials that holds promise to reduce these costs dramatically.
Numerous examples can be found in the literature demonstrating the use of sol-gel methods to synthesize hybrid inorganic-biological materials. Despite these successes, there are limitations of sol-gel based methods for long-term storage and of complex biological samples especially due, in general, to (a) processing conditions such as the presence of alcohol as a co-solvent or byproduct, and the use of acids to as catalyze silica condensation, (b) biological complexity: high salt concentrations in body fluids and complex mixture of enzymes, cells etc with varying charge, and finally (c) inorganic-biological interactions such as continuous condensation results in generation of mechanical stresses that arise at the cell or protein/silica interfaces upon encapsulation, which can limit biological activity.
In this presentation, we will discuss promising new developments and procedures for silica encapsulation of complex biological fluids that address the challenges described above. Specifically, we have developed two novel silica encapsulation techniques:(a) a chemical vapor deposition (CVD) approach and (b) a microwave-based sol-gel process.
The CVD approach involves the diffusion of a vapor of silica precursor molecules into an aqueous solution. Using this process, we have immobilized several types of complex systems including cells, enzymes, trans-membrane proteins and lipid bilayers. The microwave-based sol-gel technique that can reproducibly result in silica formation in as little as 5 minutes, which is much faster than other sol-gel techniques, for encapsulation of biological materials. We will also describe the incorporation of various poly-NIPAM materials to minimize mechanical stress at the silica interface, as well as help promote the release of biologicals from these hybrids silica gels.

Invertebrates have diverse mechanisms for the sclerotization of body structures. For example, the marine polychaete, Nereis virens, has mechanically robust mandibles which are unique due to their lack of mineralization. Nvjp-1 is a histidine-rich protein that was discovered as a component in the N. virens jaw and is proposed to be involved in the hardening of the mandible along with zinc binding. In the present study we have developed methods for the expression, purification and manipulation of Nvjp-1 which has led to the fabrication of fibers, films and bars. These materials are made using multiple cross-linking strategies followed by transition metal treatment. Mechanical testing has been done in both a hydrated as well as dry state to determine how metal binding and hydration affect the mechanical properties of Nvjp-1 based materials. Uniaxial tensile testing of the cross-linked material has shown substantial changes in mechanical properties between the two states in the presence and absence of various transition metals. Nanoindentation and rheological testing have also been employed to provide additional information about how metallization and hydration affect the mechanical properties of materials made from Nvjp-1. These results will be important in the creation of scalable, synthetic biomimetic materials that have tunable properties directed at the spatial and temporal induction of material hardening.

Surface modification of titanium implants is one of the most common practices to improve osteointegration. One of those techniques is the electrochemical treatment of titanium, called anodizing, in which the titanium surface is coated by an anodic oxide coating which is the consequence of titanium oxidation during electrode (anode) reaction. Besides to improve bone cells adhesion and corrosion resistance, being a surface treatment, it is expected that anodizing process could have some influence on mechanical properties the implants. In this work was evaluated the influence of conventional titanium anodizing in fatigue life, as well as the effect of titanium spark anodizing on contact mechanics properties. Analysis of SEM, AFM and FIB characterization, as well as the biological testing, allowed establishing the processing conditions suitable to control suitable roughness parameters for osteointegration. Comparison of fatigue life, R = 0.1, between un-anodized and anodized titanium samples showed that anodic coating improves fatigue resistance due to a combined effect of increasing both surface hardness and compressive residual stress. On the other hand, contact mechanics characterization (Hertzian and scratch testing) of spark anodized samples showed that the micro porous anodic coating exhibits a dissipations stress during contact loading associated to a re-accommodation of the porous structure. This pseudo-plastic response is the main responsible of the high contact resistance of spark anodized titanium.

Fuel-free micro/nanomotor has attracted considerable attention due to its promise for future in-vivo biomedical applications such as targeted drug delivery. Here we describe a new design and theoretical modeling of high-speed magnetically-propelled nanowire swimmers which exploit the flexibility of nanowires for propulsion. The new magnetic nanowire swimmers (5-6 mu;m in length and 200 nm in diameter) can be prepared in large scale using a simple template electrodeposition protocol. These readily prepared nanoswimmers display both high dimensional propulsion velocities (~21 mm/s) and dimensionless speeds (in body lengths per revolution) when compared with natural microorganisms and other artificial motors. They can also move efficiently in real biological environment (such as human serum). In addition, the first example of directed delivery of drug-loaded magnetic polymeric particles using such flexible nanoswimmers is also described. They are able to transport micrometer particles at high speeds of more than 10 mu;m/s (more than 0.2 body lengths per revolution in dimensionless speed). The effect of the cargo size on swimming performance is evaluated experimentally and compared to a theoretical model, emphasizing the interplay between hydrodynamic drag forces and boundary actuation. Potential applications of these cargo-towing nanoswimmers are demonstrated by using the directed delivery of drug-loaded microparticles to HeLa cancer cells in biological media. Transport of the drug carriers through a PDMS microchannel from the pick-up zone to the release microwell is further demonstrated. We expect that such magnetically driven nanoswimmers will provide a new approach for the rapid delivery of target-specific drug carriers to predetermined destinations.

Nanoscale silicate platelets (NSP), derived from the exfoliation of natural montmorillonite clays, show an unexpected antimicrobial property. A physical trapping mechanism has been proposed because the clay platelets can inhibit the growth of diverse range of bacterial species. The growth of the drug-resistant species such as methicillin-resistance Staphylococcus aureus and silver ion-resistant Escherichia coli were completely inhibited at applications of 0.3 wt% NSP. The generation of singlet oxygen species from NSP under ultraviolet irradiation and their detection using spectroscopic electron paramagnetic resonance spin trapping. The ability to generate singlet oxygen species was first observed for the clay platelets that showed a high-aspect-ratio geometric shape (ca. 80 × 80 × 1 nm) and the presence of surface ionic charges (ca. 18,000 ions/platelet). By comparison, the pristine clay with a multilayered structure failed to generate any singlet oxygen species. The ability to emit singlet oxygen species provides direct evidence for the antimicrobial ability of clay through a non-chemical mechanism, which opens the potential for medical use.

The structure, organic matrix and mineral structure of the scar (the interface between the adductor muscle and the shell) in Mytilus galloprovincialis were investigated. The scar was found to be a hierarchically multilayered structure composed of organic matrix and structural different minerals. Different from the calcite structure of the nacre, we have identified the top layer of the scar to contain structurally organized columnar aragonite. Study of the organic matrix showed that there was at least one protein that seemed to be preferentially localized in this prismatic layer. Since the scar is the most important stress distribution site in the mussel, the function of the columnar structure and the matrix protein would be discussed in relation to a similar structure at the tendon-bone connection site.

Recently, hybrid structures built using alive microorganisms as templates to obtain self-organized structures of colloidal inorganic nanoparticles have been intensively studied because their unique properties and wide technological applications. In this work, we are interested in the use of fungi as biotemplates to obtain self-assembled systems of gold nanoparticles, synthesized by the Turkevich method, forming stable mesostructures with potential use for sensors and biosensors via surface-enhanced Raman scattering. Several experimental parameters were investigated, for instance the influence of the fungi type, culture media and the concentration of gold nanoparticles. It was also analyzed the effect of heat treatment on the final microstructure and the performance of the materials as substrate for SERS. The filamentous fungi are prepared using three types of fungi (Penicillium brasilianum, Xylaria sp and Aspergillus aculeatus) that were growth in three different culture media (CZA, CZA-lev and Potato Dextrose). The filamentous fungi was prepared by adding the spores in the culture medium and keeping the system under controlled conditions for 5 days aiming the microorganism growth. After this time, the culture medium was separated, and only the fungus hyphae was used to fabricate the hybrid material. The colloidal gold nanoparticles were prepared separately through the reduction of a gold salt with citric acid at temperature near to boiling point. The hybrid material was obtained by the addition of the stable gold aqueous dispersion on the fungi hyphae previously separated. After few hours, the nanoparticles adhered to the cell wall, covering it in multiple layers resulting in uniform gold microtubules with controlled thickness, forming a reddish purple macroscopic material on the bottom of the Erlenmeyer flask that could be separated easily from the aqueous media, reducing the total concentration of nanoparticles in suspension from 5-10 days. Gold nanoparticles and microtubules were characterized by X-ray diffraction, UV-Vis spectroscopy and scanning electron microscopy (SEM). The SEM images confirmed the formation of the hybrid material, which could be visualized with naked eyes. Gold nanoparticles were observed decorating the fungal surface, forming a uniform surface on the cell wall. These structured systems showed a uniform morphology controlled thickness choosing the correct concentration of gold nanoparticles in suspension. These hybrid structures showed potential for using as substrate sensor or biosensor via surface-enhanced Raman scattering for identifying molecules of benzotiol.

It is our hypothesis that surface directed transport of chemical agents offers the potential to transport, separate, and concentrate, chemical compounds with unprecedented accuracy and speed. Here we will demonstrate the use of chemical gradients to selectively accelerate the transport of chemical compounds across a surface, leading to both chemical separation and concentration. Both the theoretical underpinnings and experimental results will be presented. Two major classes of gradients will be discussed. The first is based on the surface initiated polymerization (ATRP) of a lateral gradient formed by two different polymer brushes, and the second is based on a thin film of a hydrogel which has been locally chemically functionalized to form a gradient structure. Gradients are formed using both photopatterning strategies and localized polymerizations depending on the desired width of the gradient and the specific chemistry of interest. We find that a chemical potential gradient can accelerate the transport of a molecule many fold over simple diffusion, and that the chemical potential gradient can even be used to separate various molecules. We also hypothesize that surface chemical gradients can be used to deliver compounds of interest to cells lying on a surface.

Dental enamel is the strongest tissue in the human body with a hardness that may exceed 6 GPa, an elastic modulus that can be over 100 GPa and also high fracture toughness. Mechanically it is much stronger than typically seen in mineralized tissues such as bone, yet its main inorganic constituent is the same as other mineralized tissues in humans, namely defective hydroxyapatite, HAp. Enamel&’s strength is a result of it being over 95% HAp with a complex hierarchical structure that makes it mechanically, and to some extent chemically, very resilient. It is has the remarkable ability to withstand many years of constant wear, chemical attack and fluctuations of up to 100 °C in temperature. Here we will present the results of our on-going studies of the mechanics, structure and chemistry of dental enamel. These experimental and modeling studies have encompassed intra- and inter-tooth variations and how these are modified by acid attack associated with development of caries. The research highlights the importance of the multiscale structure of enamel in ensuring its survival even under adverse conditions. In particular the combination of nanoscale hexagonal crystals packed in an organic matrix enables dental enamel to exhibit a wide range of properties including: high hardness; high elastic modulus; toughness; mechanical anisotropy; viscoelasticity; inhomogeneous demineralization; the ability to remineralize. Valuable lessons can be learnt from the studies in terms of treating dental diseases and developing strong, biomimetic nanocomposites with resistance to environmental degradation.

Many marine invertebrates utilize chitin and melanin as a building block to form their hard tissues. Their hard tissues made of chitin and melanin containing composites are appealing prototype building blocks due to their extraordinary mechanical properties and light weight compare to the inorganic materials, correlated with energy efficiency. Here, a marine hydroid perisarc of Aglaophenia latirostris Nutting, 1900, was investigated to understand how nature design the hard tissues made of chitin and melanin containing components. Chitin and melanin were constituted 10wt %, 53 wt% of the perisarc, respectively. Interestingly, a DOPA (3, 4-dihydroxyphenylalanine) containing protein and iron was detected from the perisarc, as same as adhesive and coating of marine mussels. A resonance Raman spectrum due to DOPA-iron (III) complex formation was also detected in the chitin containing perisarc, implying the existence of DOPA-iron (III) interaction around chitin structure for the first time.

Resveratrol is a natural molecule contained in red wine. Its antioxydant properties are usefull to prevent cardio vascular diseases. Glycosylation of resveratrol protects it from enzymatic oxidation, and thus extends its lifetime in the cell. Glycosyl-resveratrol has surfactant properties. Research on mesoporous silica as drug-delivery systems has been increasing rapidly during recent years for it allows an easy storage of a large amount of drug in mesopores while the silica surfaces can be functionalized for controlled drug delivery and vectorization in a very flexible way. A system in which a same molecule both the template and the therapeutic load represents an original highly sustainable approach in this field, and avoids the expensive use of toxic or non usefull surfactants as sacrificial agents. We have developed this original approach by using glycosyl-resveratrol as a template. The resulting Hybrid silica-based materials were processed as thin- films and spray dried powders. The thin films were used as a model structure for the study of the structuration of the material, as well as the drug release properties and the degradation of the inorganic network. First, the self-assembly of the glycosyl-resveratrol was shown to mesostructured the hybrid material. The release of glycosyl-resveratrol in Phosphate Buffer Solution (PBS) was measured in situ by UV spectroscopy and lasts about 2 hours at 37°C. In parallel, spectroscopic ellipsometry was used for studying in situ the structural evolution of this hybrid thin film upon exposure to Phosphate Buffer Solution (PBS) by immersion. The fast decrease of refractive index of the thin film shows that the degradation of the silica network by hydrolysis has necessarily an impact on the drug release kinetics in this case. As a consequence, tailoring release properties of such a hybrid material is possible by tailoring the degradation kinetics of the matrix (by using binary oxide compositions or hybrid silica-based matrices for example) can be a tool for tailoring the drug release properties from a drug-templated mesostructured therapeutic vector.

Breast cancer preferentially metastasizes to bone and induces pathological remodeling. Although the exact mechanism of this process remains unclear, there has been evidence implicating that the nanoscale properties of the bone mineral may affect breast cancer metastasis. Hydroxyapatite (HA, Ca10(PO4)6(OH)2) is a calcium phosphate mineral closely related to the inorganic component of bone and often used for conferring structural and mechanical properties to bone. We have developed 2-dimensional platforms containing HA and fibronectin (FN), a major extracellular matrix (ECM) protein, to mimic the interface between bone and the ECM for the study of breast cancer metastasis. A library of three HA nanoparticles with various crystallinities and narrow size distributions were synthesized through a two-step method in which a typical wet precipitation reaction of a calcium salt with a phosphate salt was followed by hydrothermal aging for 0 and 3 days. Particles were washed with 0.15 M NH4OH, rinsed with acetone and dried at 20° C. The dried particles were either used as prepared or ground in a mortar. X-ray diffraction was used to determine particle phase (HA) and size (Scherrer analysis). Fourier Transform Infrared Spectroscopy (FTIR) was used to assess the crystallinity of the particles. The size and shape of the HA nanoparticles were determined by Transmission Electron Microscopy (TEM). Both the size and crystallinity of the particles were found to increase with longer hydrothermal aging time. Finally, Forster Resonance Energy Transfer (FRET) was used to assess FN conformation by adding traces amounts of FRET labeled FN to the 2-dimensional HA containing platforms in phosphate buffered saline. The changes in FN adsorption and conformation over time were further correlated with the specific properties of HA, such as size, shape, and crystallinity. Overall these results provide further insights into the role of materials properties of HA in controlling inorganic-organic interactions in bone.

A big challenge in the Florida phosphate mining industry is the separation of dolomite (CaMg(CO3)2) contaminants from the desired francolite mineral (a fluorapatite (Ca5(PO4)3(F OH)). In this study, phage display techniques were utilized to select phage clones with specific binding affinity to francolite, which could then be used as bio-collectors in the flotation technique for particle separation. A phage clone with a 12-mer francolite binding peptide was able to concentrate the content of francolite in a bench-top flotation process containing mixed minerals. The hydrophobicity of the phage coat proteins was evaluated using captive air bubble techniques in order to correlate with the effect of the phage coat proteins on the recovery rate of the minerals. It was found that the coat protein of M13 phages could enhance the hydrophobicity of the francolite surface. The chemical and physical characteristics of the phage peptides were evaluated to explain the selectivity of the phage, and the potential for using this biotechnology approach for a commodity industrial application is discussed.

The idea of building self-healing mechanisms in synthetic materials is very appealing and has attracted much interest over the last decade. The goal is to replicate the ability of biological structures like bone to adapt and self-repair. Much of recent research in the field has focused on the development of self-healing polymers. A common strategy is the introduction in the matrix of capsules or vascular networks containing healing agents that can be released upon fracture restoring the structural integrity. This strategy, however, does not allow further healing upon secondary crack propagation. The synthesis of organic networks able to form reversible bonds is an attractive alternative but in most cases these networks do not exhibit the mechanical properties needed for load bearing applications. An alternative is their combination with hard phases such as ceramics or carbon fibers to form structural composites. However, in these composites the organic/inorganic interface often represents the weak path for crack propagation and self-repair will depend on our ability to design self-healing interfaces.
In this work we explore the adhesion and self-healing characteristics of the organic/inorganic interface between glass and a supramolecular polymer produced by the reaction between polydimethilsiloxane (PDMS) and boric oxide. This polymer exhibits a highly interconnected tridimensional network with a self-healing character due to the presence of reversible hydrogen bonds. It cannot act as a structural material on its own but is a promising candidate for the formation of soft interlayers between inorganic components in composites.
We use Double Cantilever Beam (DCB) test to study the adhesion and self healing response of the interfaces. The polymer exhibits a shear thickening behavior and the fracture energy shows a marked dependence with the displacement rates, varying up to two orders of magnitude when the rates change between 0.5 to 50 mm/min. Upon stopping the delamination tests two main self-healing mechanisms with very different kinetics can be observed. The first, faster mechanism, consists on the re-formation of hydrogen bonds and the second involves the spreading of the polymer to eliminate porosity and fully cover the un-bonded areas creating a continuous interface. The initial reformation of the hydrogen bonds results in a recovery of ~40% of the bonding strength; full recovery is achieved after healing for ~ 3 days. These results provide information on the adhesion and self-healing mechanisms that will be used to design intrinsically self-healing composites with improved properties.

We present a systematic design study and adhesion analysis of gecko-like adhesives matching closely the inspiring example of the gecko in dimensions and elastic modulus. Applying 3D direct laser writing (DLW) we designed and fabricated artificial hierarchical gecko-type structures on the micro- and nanometer scale [1].
The famous ability of geckos to climb walls and ceilings originates from setae, delicate hairs covering their toes. These setae consist out of beta-keratine, a stiff material with elastic modulus of around 1 - 4 GPa. Due to their hierarchical design, they are very compliant and can achieve a very high real contact area even to rough substrates. Thus, the van der Waals forces cause very high adhesion enabling geckos to stick to nearly every surface. Mimicking the gecko toes is, therefore, pursued by many research groups all over the world. However, such structures are mostly based on soft materials that have some drawbacks like degradation or rapid contamination. To overcome these issues, we used 3D direct laser writing to replicate gecko-type adhesives out of a stiff material.
3D direct laser writing is a very flexible rapid prototyping method allowing the fabrication of arbitrary nanostructures. Since the parameters of the fabricated structures can be easily varied this technique is perfect for design studies of dry adhesives. Measuring the adhesion forces by atomic force microscopy, the influence of several design parameters like density, aspect ratio, and tip-shape on dry adhesion performance were systematically examined. The interpretation of the measured adhesion was supported by adhesion maps obtained with colloid AFM tips. In this way, we demonstrate the positive impact of mushroom-shaped tips for stiff materials with lateral dimensions in the nanometer range. In addition, we confirmed that hierarchy is favorable for artificial gecko-inspired dry adhesives.
[1] M. Röhrig, M. Thiel, M. Worgull, H. Hölscher: 3D Direct Laser Writing of Nano- and Microstructured Hierarchical Gecko-Mimicking Surfaces. Small 8, 3009 (2012).

The exoskeleton of crustaceans is formed by the cuticle, a continuous tissue secreted by the epithelial cells that covers the whole body of the organisms. On the molecular level, the cuticle consists of chitin and proteins that form fibrils, which are hierarchically organized over several levels and can be associated with different biominerals, mainly carbonates and phosphates. On the higher levels of hierarchy, the cuticle forms skeletal elements with elaborate functions. The physical properties of skeletal elements are adapted to these functions and can be very diverse, like providing mechanical stability in the shell of most body segments, elasticity in arthrodial membranes, or wear resistance, friction reduction and sliding ability in joint structures and mandibles. The required properties for each skeletal element are adjusted by local modifications in structure and composition. The function of the exoskeleton as a whole is ensured by the formation of transitions between parts with different physical properties. These transitions emerge from structural interfaces that are generated on different hierarchical levels.
We investigated structure-composition-property relationships of functionally differentiated cuticle parts: mandibles of selected Crustacea species with different feeding habits and transitions between load-bearing cuticle and arthrodial membranes, with particular attention to the interfaces between highly mineralized and unmineralized cuticle parts. The microstructure was characterized using light (LM) and scanning electron microscopy (SEM). The local chemical composition was investigated using energy dispersive X-ray spectroscopy (EDX) and confocal µ-Raman spectroscopy. Nanoindentation tests were performed to study the local stiffness and hardness.
The experimental data are used as input for a hierarchical model for cuticle that uses ab initio calculations for the properties of basic components like chitin and calcite and hierarchical homogenization performed in a bottom-up order for the higher hierarchical levels. We developed a hierarchical model for the elasto-viscoplastic cuticle properties at large deformations using a Fast Fourier Transforms (FFT) approach that is able to describe the local development of stress and strain fields within the material, including those at the interfaces.
Understanding the nature of these interfaces can provide inspiration for the development of multifunctional composite materials with seamless transitions between parts with different properties.

In the field of drug delivery there is a growing need for biocompatible drug carriers. For example, medical implants such as orthopedic, dental, and vascular stents, may require subsequent drug therapy regiments to prevent infection or decrease inflammation. Release of bioactive agents from the implant surface rather than systemically can improve patient rehabilitation. In the present work biocompatible TiO2 nanotube- decorated implants loaded with bioactive agets are developed as a more suitable alternative route toward drug-eluting implants. The novel anodization technique without the usage of any harmful chemicals ensures the robustness and biocompatibility of the implants. The results demonstrate an increase in the degree of osseointegration compared to conventional titanium surfaces. An anti-inflammatory drug (Sodium Naproxen) was encapsulated inside these biocompatible TiO2 nanotubes by means of self-sustained diffusion, which allows one to avoid any high temperature or pressure procedures. Such drug-intercalated nanotubes can be used as stand-alone drug carriers or as a surface modification of orthopedic and/or dental implants. The method of preparing biocompatible TiO2 drug carriers can also be extended to any other bioactive systems, thus eliminating completely the chances for cytotoxicity and avoiding high energy requirements of the other existing methods of drug encapsulation and delivery.

The number of percutaneous coronary intervention (PCI) procedures, commonly known as balloon angioplasty, has increased by 30% over the past 10 years totaling more than 1.3 million patients in the U.S. annually (1). The large increase in procedures is due to a rise in heart disease as well as technological advancements, which have led to safer, more effective practice. Still, PCI procedures are not without post-procedure complications including thrombosis and restenosis (2). PCI procedures result in injury of the vessel wall, which in turn initiates inflammation and coagulation through platelet activation. Attenuating the inflammatory and coagulation responses can mitigate the negative impact of the injury (2-4).
Drug-eluting stents (DESs) have been used widely due to their ability to prevent restenosis. However, complications of DESs are development of late in-stent thrombosis and the need for long-term systemic anticoagulation. The DESs used in clinical applications deliver an anti-proliferative agent, such paclitaxel (PTX), which inhibits not only smooth muscle cells (SMCs) proliferation, but also prevents proliferation of endothelial cells (ECs). Controlling the late thrombosis, which occurs between 1 month and 1 year after stenting, requires solving fundamental problems related to the lack of EC growth in the stented area, while maintaining the inhibition of intimal hyperplasia offered by the antiproliferative drug. Thus, to traverse the complications associated with PCI, the investigation of therapeutic delivery has become an integral topic in modern research.
Our work investigates the hypothesis that the late in-stent thrombosis is largely due to the lack of growth of ECs over the stented area, and further, that both SMC proliferation and lack of EC growth can be controlled through early steps of preventing initial platelet binding, attenuating inflammation and limiting the SMC mitogenic response. We have investigated a group of technologies including an in vitro culture system that provides the opportunity to evaluate therapeutic response in contractile and proliferative vessel models, development of quick-dissolving coatings for rapid, local therapeutic delivery form stents or balloons, development of bioinspired nanoparticles for additional controlled local delivery of therapeutics, and development of bioactive therapeutics. These technologies synergistically improve outcomes following balloon angioplasty.
1. Lloyd-Jones, D., et al., Circulation 2009, 119, (2).
2. Byrne, R. A., et al., Drug Safety 2009, 32, (9), 749-770.
3. Danenberg, H. D., et al., Circulation 2002, 106, (5), 599-605.
4. Tanguay, J. F., et al., Thrombosis and Haemostasis 2004, 91, (6), 1186-1193.

Bone tissue damages due to traumas and degenerative diseases are potentially treatable by tissue engineering practice. Remarkable surface and bulk properties of titanium are useful because they can be suitably tailored to mimic bone tissue, as can be done by producing porous materials by techniques like powder metallurgy (PM). Surface of porous titanium implants is an important factor in wound healing process because of it is going to modify cells growth. The aim of this (study was to compare osteoblastic cell behavior on different titanium implants surfaces (1250 C y 2h): PM loose sintering (18% of porosity) and space holder with ammonium bicarbonate (50%, 800 MPa). Biological characterization was performed through in vitro assays with MC3T3-E1cell line (mouse pre-osteoblast, CRL 2593) at a cell density of 10000 cells per sample at 3 different times, 24h 48h and 72h. Cell proliferation assays were carried out by using AlamarBlue® in order to evaluate cell adhesion and proliferation on modified titanium, throughout fluorescent measurement. Total cellular protein content and ALP activity served to determine cell proliferation on samples. Scanning electron microscopy and Actin staining was performed to evaluate surface and cytoskeletal organization of cell seeded samples at different times. Biological results indicate that modified titanium allowed cell adhesion and cell proliferation by different in vitro tests. Actin staining microscopy permitted the visualization of cell morphology during cell proliferation processes. Through SEM images it was possible to observe of fully cell-covered modified titanium after seeding MC3T3 cell line and cultured during 3 days. Some pores were totally covered at 72h by flattened cells. The main findings of this work confirm what have been previously reported by the authors about the influence of both porous curvature and inner roughness on the osteoblast behavior inside the pores.

Surgical lenses in laparoscopes, and arthroscopes fog during surgery, increasing its duration by up to 40%, and infection rates and scarring due to exposure to air from repeated scopes withdrawal for cleaning. Modeling nucleation of fluids on surfaces yields insight that they need to condense in 2-D films for transparency rather than 3-D droplets where refraction at multiple gas/fluid interfaces lead to the opacity referred as “fogging”. Our VitreOxtrade; model enables for control of water affinity of lenses made either from bio-compatible polymers, and boro-silicate using a nano-scale molecular mesh applied as a bio-identical visco-elastic colloidal emulsion. The VitreOxtrade; model [1-5] meets a 100% success rate in the lab over in over 300 experiments. Twenty surgical trials in the OR yield a success rate of 90 %, with loss of vision due to the presence of blood or blood proteins, not fogging.
We studied the common blood protein, heparin, which prevents coagulation, within the VitreOxtrade; model using applications of colloidal emulsions. Heparin behaves like H2O on hydrophobic surfaces. It does not prevent fogging nor interfere with Vitreoxtrade;. Next, we investigated fibrinogen as agonist agent because it causes coagulation. Fibrinogen applied to various surfaces in combination with VitreOxtrade; can prevent not only fogging but blood wetting. Our research shows that proteins combined with the correct colloids emulsion can help avoid whole blood accumulation on lenses, and leads to a new model, ProteinKnoxtrade;.

[1] U. S. Patents Pending (2008, 2009,2 010, 2011, 2012)
[2] PhD Dissertation, Q. Xing, ASU (2011).
[3] N. Herbots, et al. NIMB B (2011).

Titanium and some of its alloys are still the best metallic biomaterials for bone replacement. It is also known that despite their well-known advantages for this application, surface properties of implants and prostheses can be improved in order to obtain better fixation and tissue growth. Control of microtopography is an accepted factor which, within a certain specific range, has shown to have positive effects in osteoblast adhesion and in vivo osteointegration. However, the influence of both nanotopography and nanostructuring of titanium in cellular adhesion and tissue generation is still an issue that has to be addressed by the biomaterials scientific community. Recent work of the authors have demonstrated the capability of Directed Irradiation Synthesis (DIS) techniques to drive initial micro-rough or micro-structured metallic surfaces (Si, Au, Pd) to both nanoscale kinetic roughening and nanostructured surfaces, depending on the irradiation parameters. In this set of experiments, samples of commercially pure titanium, CP Ti, and Ti6Al4V alloy were surface modified by different conditions of DIS and analyzed for their subsequent structural, chemical, and biological properties. Analysis of SEM, AFM and XPS characterization established the processing conditions suitable to control the nano-roughness of both CP Ti and Ti6Al4V samples. Only extremely controlled DIS conditions of ion energies, fluences, and angles of incidence were suitable for partial nano-patterning of both kinds of titanium surfaces. Nanorough surfaces obtained by DIS in addition to certain surface curvature of Ti samples have shown improved osteoblasts adhesion. With respect to bone tissue growth stimulation from nano-patterned Ti surfaces, preliminary cell culture results indicate that more studies have to be developed in order to confirm repeatability of cellular growth and behavior.

Stenting is an important procedure in many medical interventions, including vascular, esophageal, and ureteral therapies. Permanent stents, whether bare or drug-eluting, have the potential to cause late stent thrombosis [1,2], which can ultimately lead to myocardial infarction and death of the patient. Permanent stents are also problematic when inserted into a blood vessel of a growing child, as the vessel may ultimately grow too large for the stent [3]. Resorbable metal stent designs, such as pure iron (Fe) and magnesium (Mg), thus far have not been feasible due to poor elasticity and strength, while polymer designs require large stent struts, which increase the overall hemodynamic impact of the implant. We present bioabsorbable metal wires comprising high-purity iron (Fe), manganese (Mn), magnesium (Mn), and zinc (Zn), as candidate materials for disappearing stent designs. Preliminary mechanical testing on these materials showed yield strengths exceeding 1 GPa and elastic strains of greater than 1% [4]. In this study, we compare the thrombogenic properties of these materials with 316L stainless steel, as a low-impact control, and copper-coated wires for a thrombogenic control. Venous whole blood from a single pig was pumped through each stent for 1 hour. The whole blood was counted for red cells before and after exposure, and the plasma from each sample was assayed for beta-thromboglobulin and thrombin-antithrombin complex to determine the thrombogenic impact of each material. The stents were then imaged for cell adhesion with scanning electron microscopy (SEM). The results suggest that iron manganese stents are similar to 316L in terms of thrombogenicity. We extend this study to investigate ion-beam modification of the stent surfaces to alter the physiological response.
[1] McFadden, E.P. et al. (2004) “Late thrombosis in drug-eluting coronary stents after discontinuation of antiplatelet therapy.” Lancet 2004(364) 1519-1521.
[2] Waksman, Ron. (2006) “Update on bioabsorbable stents: from bench to clinical.” Journal of Interventional Cardiology 19(5) 414-421.
[3] Peuster, M. et al. (2001) “A novel approach to tempoaray stenting: degradable cardiovascular stents produced from corrodible metal—results 6-18 months after implantation into New Zealand white rabbits.” Heart 86(5) 563-569.
4] Schaffer, Jeremy E., Eric A. Nauman, and Lia A. Stanciu. (2012) “Cold-drawn bioabsorbable ferrous and ferrous composite wires: an evaluation of mechanical strength and fatigue durability.” Metall and Materi Trans B 43(4) 984-994.

Novel multilayer capsules have been prepared using the Layer-by-Layer (LbL) by combining the excellent properties of novel 2-dimensional material ‘graphene oxide&’ (GO) with polymers for biomedical applications. Here, we describe a novel strategy for the fabrication of near-infra red (NIR) light responsive composite multilayer capsules by introducing the GO into the LbL assembly without addition of other NIR-absorbing additives such as metal nanoparticles (NPs) and carbon nanotubes (CNTs) into the shells of the capsules. The GO plays a dual role; it serves as a structural component of the capsules, as well as, a strong NIR-light absorbing agent. The laser illumination was performed by irradiating the capsules with NIR pulsed laser at 1064 nm wavelength. (Nd:YAG laser ~10 ns, 30 mW, 10 Hz). The rupturing of the capsules and changes in morphology of the capsules were analyzed using optical microscopy and electron microscopy. The GO composite capsules were ruptured in ‘point-wise opening&’ manner due to local heating phenomenon caused by irradiation of the GO by NIR-laser. The Raman spectrum analysis of laser treated GO composite capsules indicates that GO remains structurally intact after 1064 nm pulsed laser irradiation. We have demonstrated successfully the encapsulation of anticancer drug doxorubicin into these capsulses and its subsequent release by irradiation with a 1064 nm pulsed laser.
References
1. Rajendra Kurapati, Ashok M Raichur, Chem. Commun. 2012, 48 (48), 6013-6015.
2. Rajendra Kurapati, Ashok M Raichur, Near-Infrared Light-Triggered Graphene Oxide Composite Microcapsules: A Novel Route for Remote Controlled Drug Release, (Submitted to Chem. Commun.).

Hybrid inorganic-biological materials with combined physiochemical and biological properties have attracted much attention for emerging applications including medical, diagnostics, catalysis, biofuel cells and nano-devices. Inorganic nanomaterials impart unique optical, electrochemical and magnetic characteristic; while biological materials provides specific bio-recognition capability, and offer excellent biocompatibility. Integration of various inorganic and biological materials into a one entity allows intimate contact and enhances the synergistic interactions between the hybrid materials. A simple and flexible fabrication technology enables precise manipulation of physiochemical properties and biological function is the key to achieve practical applications. Here, we report a facile method to fabricate multifunctional hybrid inorganic-biological colloidal particles using calcium carbonate template. The fabrication procedure is simple that only required simple mixing and centrifuge step in aqueous medium at physiological pH and room temperature with a single self-assembly step. Various hybrid inorganic-biological particles with optical, electrochemical and magnetic properties in combination with bio-catalysis and bio-recognition properties have been fabricated for emerging applications such as bead-based diagnostic and bioseparation. The physiochemical and biological properties of these hybrid inorganic-biological particles were well characterized. Our technology provides a simple and cost-effective fabrication process to design and fabricate hybrid inorganic-biological colloidal particle with tailored functions in a precise and controllable manner. Advances in fabrication technology are creating new commercial opportunities in high value added industries such as pharmaceutical, diagnostic, medicine and bio-electronic devices. Our long-term goal is to bring such multifunctional hybrid inorganic-biological colloidal materials from research to industrial applications and eventually making them to the “Swiss Knife of particles”.

Hydrogels are 3-dimensional polymeric networks with water content as much as over 99 wt%. They are utilized in various applications including tissue adhesives, extracellular matrices for tissue engineering and repair, drug delivery vehicles, and actuators. However, hydrogels are fragile and have a very narrow elastic range, which makes them unsuitable for most load-bearing situations. Nanocomposite hydrogels, composed of inorganic nanoparticles dispersed within a crosslinked organic polymer network, have demonstrated increased materials properties over conventional hydrogels. These materials rely on weak physical interactions (e.g., hydrogen bonding, ionic interaction, etc.) between the inorganic and organic components to achieve elevated mechanical properties as well as the ability to recover after repeated deformation. Chemically crosslinked polyacrylamide (PAAm) nanocomposite hydrogels were prepared with inorganic nano-silicate, Laponite, and dopamine methacrylamide (DMA). DMA consists of a biomimetic adhesive side chain found in mussel adhesive proteins, covalently linked to a polymerizable methacrylate monomer. Copolymerizing DMA into a PAAm hydrogel strongly enhanced the interfacial interaction between the polymer network and Laponite. Nanocomposite hydrogels demonstrated reduced water content and increased materials properties that were dependent on both the Laponite and DMA contents. Nanocomposite hydrogels with relatively low DMA and Laponite contents (2-3 wt% for each) demonstrated maximum compressive stress, Young&’s modulus, toughness, and storage and loss moduli values that were over an order of magnitude higher than control gels. DMA-containing nanocomposite hydrogels also demonstrated improved fracture resistance to compressive loading, capable of repeated compressed to 80% strain without rest while exhibiting compressive stress of over 1 MPa. The catechol side chain of DMA likely formed strong reversible bonds with Laponite, which can be repeatedly broken and reform to dissipate fracture energy while minimizing permanent damage to the network architecture. An injectable tissue adhesive was formulated with a 4-armed polyethylene glycol (PEG) end-functionalized with dopamine. Increasing Laponite content increased lap shear adhesive strength using wetted pericardium tissue as the biological substrate. Incrased adhesive strength was presumably due to enhanced bulk cohesive properties of the nanocomposite hydrogel.

Nanoscale work may have been a major energy source for the origin of life. If life originated between muscovite mica sheets (1), then the open-and-shut movements of these sheets would have provided an endless source of mechanical energy for doing work on the molecules between the sheets. The open-and-shut movements could have been caused by fluid flow or temperature cycles. The mechanical energy would have been available for both chemical reactions and macromolecular assembly processes. A force of 1 piconewton moving a distance of 1 Angstrom is equal to the energy of a peptide bond, which has free energy of hydrolysis of -2.4 kcal/mole. The earliest life forms may have been organic-inorganic hybrids, because the surfaces of muscovite or other clay minerals may have been the site where life originated. To test the muscovite hypothesis for life's origins, one needs to devise ways of making mica sheets move open-and-shut in solutions containing plausible prebiotic reaction mixtures. Two approaches for doing this are, first, using an atomic force microscope (AFM) to move mica sheets open and shut and, second, using thermal cycling, as in the polymerase chain reaction (PCR).
(1) HG Hansma, J Theor Biol 266:175 2010

Exploiting robust, efficient, and orthogonal chemistry to synthesize macromolecules of well-defined architecture and chemical functionality has widened the scope of polymer systems in modern technological applications. The fabrication of sub-100 nm features with bioactive molecules is a laborious and expensive process. To overcome these limitations, we present a modular strategy to create nanopatterned substrates (ca. 25 nm features) using functional block copolymers (BCPs) to controllably promote or inhibit cell adhesion. A single type of BCP was functionalized with a peptide, a perfluorinated moiety, and both compounds, in order to tune nanoscale phase separation and interactions with fibroblast cells and human mesenchymal stem cells. The focal adhesion formation and morphology of the cells were observed to vary dramatically according to the functionality presented on the surface of the synthetic substrate. It is envisioned that these materials will be useful as substrates that mimic the extracellular matrix (ECM) given that the adhesion receptors of cells can recognize clustered motifs as small as 10 nm, and their spatial orientation can influence cellular responses.

Cocoon silk has been exploited for centuries due to its biocompatibility, low allergic response, and high production yield. Especially, spider silk has got much attention for half centuries due to its superior mechanical properties. Nevertheless, practical use of spider silk has been restricted due to limited amount of availability. Therefore, from 1960s, scientists have studied fundamental sequential and structural information on silk and attempted to realize artificial spider silk. Particularly, despite tremendous opportunity to discover new silks with extraordinary properties, scientists have only focused on certain spider and cocoon silks. For example, dragline, which is well known as the strongest spider silk was characterized mainly confined to Nephila clavipes. However, in recent years, there is a report that Caerostris darwini&’s dragline shows the best toughness ever as well as almost twice higher of elasticity than previous ones. In addition, new types of silk or silk-like proteins from spider, bee, ant, beetle, mussel, pearl oyster and shrimp have been introduced. Considering infinite potential of excellent silk's existence, discovery of new silks from unrevealed Nature and to fabricate them for industrial usage will be an optimum way to acquire the original technology at the first in the world and to proceed forward as a pioneer in sea anemone silk-based biomaterial. Here, we propose a new silk-like protein which has high contents of glycine and proline with decamer repeats derived from a tiny starlet sea anemone (Nematostella vectensis). As sea anemone varies their length and width almost ten-fold by shrinking rapidly and expanding itself under stimulus, we assumed that certain peculiar protein, alike silk, along with their skin and tentacles plays an important role in contraction and relaxation, and its existence was confirmed by immunohistochemistry method. With this observation, we launched recombinant sea anemone protein production in Escherichia coli system and named it as ‘aneroin&’. As previous silk studies have some problems on expression of silk proteins in diverse systems, we inserted expression inducer ahead of aneroin gene and optimized genetic codons suitable for E. coli. With wet spinning, micro-scale silk fibers were smoothly spun with no defects, and nano-scale silk fibers also successfully spun using electro-spinning method. After wet spinning, post-treatment, drawing, and tensile test, we analyzed mechanical properties (strength, stiffness, extensibility, and toughness) of aneroin silk fiber and compared with other biomaterials including recombinant spider silks.

Gold is known for its excellent biocompatibity because of its inert chemical activity, and is thus widely used in biological applications. Surface of gold can also be modified by depositing alkanethiols to form self-assembled monolayer (SAM). Furthremore, by altering the chemical composition of SAM, the surface properties of gold can be easily tailored. It is well known that surface properties strongly affect the behavior of cell adhesion and proliferation because it is required for eukaryotic cells to adhere through the extra-cellular matrix. Using a series ratio of amine and carboxylic acid functional group on SAMs that yields positively and negatively charged surface, respectively, surfaces with a series zeta potential ranging between +50 and -150 mV are obtained at pH 7.4. Furthermore, it is found that amine and carboxylic acid SAMs on gold poses low cytotoxicity. Thus, this binary-SAM modified gold is ideal for studying the effect of surface potential on cell behaviors including adhesion and proliferation. The surface chemical composition of binary SAM is quantified using a x-ray photoelectron spectroscopy (XPS) and the actual zeta potential is measured with an electro-kinetic analyzer. NIH3T3 cells are chosen to culture on these binary- SAM modified Au and incubated for 24h. The coverage and shape of cells adhered on surfaces of various potential are examined by fluorescence optical microscopy and scanning electron microscopy.

Hydrogels have a wide range of application in the field of biomedical engineering such as protein, nucleotides and cell delivery system. Hydrophilic environment provided by hydrogel is known to be excellent condition for cell viability and proteins without denaturation. Porosity of the hydrogels can be controlled by changing the polymer concentration and by preparation condition. Hydrogels can be formed at a low monomer concentration, which is too low to form hydrogels at room temperature reaction, using cryogel method. Previous research by Harvard group reported that macroporous hydrogels containing iron oxide nanoparticles were used as a delivery carrier of stem cells, displaying a controlled release of the cells under the application of magnetic field. Inspired by the work, ferrogels were prepared and its potential role as a stem cell scaffold was investigated.
In this study, porous hydrogel containing magnetic nanoparticles was prepared by polymerizing acrylamide and N,N&’-methylenediacrylamide monomers in the presence of poly(N-vinyl-2-pyrrolidone) (PVP) coated nano-sized iron oxide with ammonium persulfate and N,N,N&’,N&’-tetramethylethylenediamine initiating system. Gelation was obtained by cryogel method under the freezing condition around -20 oC and PVP coated iron oxide nanoparticles were homogeneously entrapped in the porous hydrogel matrix. Freeze-and-thaw produced macroporous ferrogel. Acrylamide and N,N&’-methylenediacrylamide were first polymerized to form hydrogels at 2 wt% monomer concentration solution. Macroporous structure was confirmed by Scanning Electron Microscope and the average size of the pore was around 200 micrometer. Encapsulation of iron oxide nanoparticles changed the physical property of the hydrogel and the proper gelation was achieved at the monomer concentration of 4 wt% solution. Concentration of monomer affected the morphology of the gel pores, but the change was not so noticeable to claim that the prepared ferrogel exhibited macroporous structures. The ferrogel was shrinked and expanded under the magnetic field and the extents of shrinkage and expansion was strongly dependent on the magnitude of applied magnetic field. Macroporous hydrogels were reported to display larger amount of dimensional change than nanoporous hydrogels and the ferrogel in this study followed the same trend. Prepared macroporous ferrogels have a potential candidate as a scaffold for stem cell differenciation toward a specific cell lines.

We have investigated the role that quantum rod (QR) morphology, composition, and surface chemistry play in resonance energy transfer phenomena. By fine-tuning the synthetic conditions, it is possible to tailor the aspect ratio of CdSe/CdS core-shell QRs from 2-8. The aspect ratio, as well as initial core diameter, determines the spectral properties of the QR, and the emission was tuned between 600 - 700 nm. These QR were then studied in resonance energy transfer studies with molecular fluorophores, fluorescent proteins, and bioluminescent enzymes. In order to accomplish functionalization, the biomaterials were attached to the QR interface via the N-terminus histag. The results indicate that the QR are ideal candidates for resonance energy transfer, due in large part to the increased stoichiometry provided by the rod morphology, and better spectral matching. The results also indicate that core location within the QR is particularly important when interpreting the energy transfer.

Enormous uses of fossil fuels have produced carbon dioxide massively into the air, thus resulted in global warming, but the level of carbon dioxide in the air is still increasing as times go. Therefore, there have been many efforts to reduce carbon dioxide by such as reducing the use of fossil fuels, finding alternative energy sources, and capturing carbon dioxide. Herein, we aimed to reduce carbon dioxide biologically as well as producing valuable products from converting carbon dioxide to carbonate minerals. In order to this, we used recombinant carbonic anhydrase, which has been known to beable to speed up converting aqueous carbon dioxide to bicarbonate (ex. ideally 107 times), originated from Neisseria gonorrhoeae. Recombinant carbonic anhydrase was highly expressed as soluble form and we found that purified carbonic anhydrase showed comparably CO2 hydration activity to commercial BCA and significantly promoted formation of CaCO3 compounds in the presence of Ca2+ through the acceleration of CO2 hydration rate. In addition, for the practical use of recombinant carbonic anhydrase, we constructed whole cell biocatalytic system by translocating recombinant carbonic anhydrase into periplasmic space. This whole cell system showed higher stability in high temperature as well as actual plant condition. The use of carbonic anhydrase described here will be leveraged in the future for rapid reducing of carbon dioxide and making valuable product from it.

Energy transfer using nanoparticles has been studied intensively during the last decades. However, there is currently little research focusing on energy transfer using luminescent gold nanoclusters (AuNCs). Here we report the use of near infrared luminescent (~820 nm) AuNCs as energy transfer acceptors and investigate the energy transfer efficiency from several donors with different photophysical characteristics, including widely varying luminescence lifetimes. First, we synthesized bright and stable AuNCs in water using bidentate dithiol-appended poly(ethyleneglycol) (PEG) and characterized the physicochemical properties of AuNCs. The diameter of AuNCs was ~1.5 nm and the quantum yield approached ~6 % in water, which is much higher than traditional NIR dyes (Indocyanine Green, < 1% in water). The lifetime of AuNCs exhibited a bi-exponential decay with time constants of ~1.4 µsec and ~0.2 µsec. As synthesized, the AuNCs showed excellent colloidal stability over a period of years due to the water solubility of PEG and the stability of the bidentate ligand interaction with the gold surface. We used the PEG-modified luminescent AuNCs as energy transfer acceptors, pairing them with several different donor molecules including: Cy3 and Rhodamine dyes, semiconductor quantum dots, a ruthenium metal complex, and a long-lifetime terbium chelate. Critically, these vary in fluorescence lifetime from nanoseconds to milliseconds. Energy transfer efficiencies were calculated based on the quenching of the donors and the sensitization of AuNCs luminescence using both steady-state and time-resolved fluorescence spectroscopy. We observed complex quenching of the donor luminescence in the presence of the AuNCs, which may be due to a combination of FRET and proximal quenching from larger-sized AuNCs present in the size distribution. We found almost no sensitization from the shorter lifetime donors while enhanced energy transfer efficiency between the AuNCs and the donors with longer luminescence lifetimes was consistently observed.

Complex materials with micron-scale dimensions and nanometre-scale feature resolution
created via engineered DNA self-assembly represent an important new class of soft matter.
These assemblies are increasingly being exploited as templates for the programmed assembly of functional inorganic materials that have not conventionally lent themselves to organization by molecular recognition processes. The current challenge is to apply these bioinspired DNA templates toward the fabrication of composite materials for use in electronics, photonics, and other fields of technology. A novel method is presented for
integration of DNA templated structures into functional composite nanomaterials, particularly,
organization of preformed nanoparticles and metallization procedures. DNA origami templates have been modified to have DNA binding sites with a uniqueley coded sequence. Gold nanoparticles functionalized with the cDNA sequence were then attched and, then, adsorbed on silicon oxide substrates. These seed nanoparticles were later enlarged, and even fused, by electroless deposition of silver. Using this method, a variety of metallic nanostructures have been constructed.

Titanium alloys are commonly used as structural implant materials, but deficiencies regarding the amount of tissue in-growth and surface cell adhesion can limit the lifetime of the implant. Factors such as roughness, porosity, surface charge, hydrophobicity, and surface chemistry will affect the body&’s immune response to the implant as well as the ability of cells to adhere and proliferate. Stronger tissue in-growth with the surface of the implant will lead to less slippage and mechanical wear. In this study, the conductive polymer Poly (3,4- ethylenedioxy thiophene) [PEDOT] was electropolymerized on the surface of Ti4Al6Vwith biopolymers such as chondroitin sulfate to evaluate the effect of the conductive polymer with regards to cell signaling, attachment, proliferation, and tissue in-growth. Surface properties such as roughness, charge, and porosity were varied by incorporating different electropolymerization parameters. Different biopolymers were chosen as PEDOT counter ions to evaluate degradation rate, biocompatibility, and the effect of proteoglycan interaction with osteoblasts. Surfaces were characterized by FTIR, SEM and EDS. Strength of adhesion of the coatings and their ability to withstand physiological conditions were also evaluated.

Peptoids are sequence-specific oligomers composed of N-substituted glycine monomer units. Their solid phase assembly enables the exploration of sequence-structure-function relationships in ways that may recapitulate the characteristics of biopolymers. A variety of strategies have been established to organize well-defined peptoid secondary structures. Recent studies have highlighted several intriguing materials attributes of peptoids. Peptoid macrocycles, for example, can form stacked supramolecular assemblies in the solid state that resemble peptide nanotubes. We observe that the cylindrical arrays are capable of reversibly sequestering and releasing water molecules through single-crystal-to-single-crystal transformations. In addition, we have identified a library of peptoid oligomers that function as mimics of antifreeze peptides and exhibit “dual-action” activity as exemplified by ice crystal growth inhibition concomitant with melting temperature reduction. The prospects for further development of bio-inspired materials in more complex peptoid structures will be evaluated.

We will present our recent work to produce inorganic and hybrid materials at the surface of single cells and multicellular tissues, using the cell itself as a template. Our goal is to create material properties such as mesoporosity and hierarchical structure to enable function such as precision size exclusion and durability for applications in regenerative medicine, tissue engineering, and animal models of disease. The work is inspired by both natural living systems, e.g. diatoms, and synthetic materials produced in abiotic conditions. To achieve our goal we must produce the materials under conditions that are compatible with living cells and culture conditions. Sol-gel synthesis methods provide a good starting point given the now 2 decades of work doping biomolecules and cells in sol-gel produced bulk silica. Recently, we have shown that sol-gel synthesis reactions occur preferentially at the cell surface prior to bulk gelation. This finding provides an opportunity to grow thin and thick films at the cell and tissue surface by simply controlling the time the cells are exposed to a cell culture media that is saturated in silica precursors. We have characterized the resulting biosilica coatings on a range of cell and tissues include: human & mouse pancreatic islets, pluripotent mouse stem-like cells, beta cells, neurospheres, and bacterial biofilms. Material morphology and composition were measured by scanning electron microscopy and energy dispersive X-ray spectroscopy. Biological properties were measured by confocal fluorescence microscopy, live/dead staining, metabolic flux, RT-PCR, and hormone release assays. Living cell / biosilica constructs were analyzed for up to one month. Under certain cell culture conditions, the biosilica can induce secondary biomineralization resulting in a calcium phosphate coating. This approach combined with advances in synthetic biology and stem cell biology offer exciting opportunities to create new materials and engineered tissues.

Nipah virus (NiV), a highly pathogenic member of the Paramyxoviridae family depicted in the 2011 film, Contagion, is classified as a BSL-4 agent due to its numerous routes of transmission and the high mortality rates associated with infection. Despite recent advances in understanding the cellular tropism of NiV, however, treatment remains primarily supportive. To this end, we have developed mesoporous silica nanoparticle-supported lipid bilayers (‘protocells&’ - see Nature Materials (2011) 10: 389-397) that specifically deliver high concentrations of therapeutic nucleic acids to host cells that are either stably transfected with NiV genes or pre-infected with a NiV pseudovirus. To generate nucleic acid-loaded protocells, we first soaked mesoporous silica nanoparticles (MSNPs) 100-300 nm in diameter with 5-25 nm pores in a solution of siRNA or minicircle DNA designed to silence expression of NiV nucleocapsid protein (NiV-N) or matrix protein (NiV-M); due to its high surface area, the MSNP core can be loaded with high concentrations of siRNA (30 wt%) and DNA (5 wt%). Liposome fusion to nucleic acid-loaded cores results in a supported lipid bilayer (SLB) that promotes long-term (>1 month) cargo retention and provides a fluid interface for ligand display. To generate targeted protocells, we employed phage display to identify peptides that bind to human ephrin B2 (EB2), the primary cellular receptor for NiV. TGAILHP (specific clone-1, or SC1) was the predominant sequence after five rounds of positive selection against CHO-K1 cells stably transfected with EB2 and four rounds of subtractive selection against parental CHO-K1 cells. We have found that SC1-targeted protocells have a nanomolar affinity (K_d = 0.5-20 nM) for EB2-positive cells (CHO-K1/EB2, HEK 293) at both high (1.50 wt%) and low (0.015 wt%) valencies and, when co-modified with a peptide (R8) known to trigger macropinocytosis, are rapidly internalized by HEK 293 but not by EB2-negative cells (CHO-K1, PK-13). Using cells that were either stably transfected with NiV-N or M or pre-infected with a BSL-2 pseudovirus that encodes NiV-N or M, we have demonstrated that SC1-targeted, siRNA-loaded protocells silence 98% of NiV-N or M mRNA in HEK 293 at a siRNA concentration of just 30 pM without affecting NiV-N or M levels in CHO-K1. Additionally, SC1-targeted protocells loaded with minicircle DNA vectors that encode NiV-N or M-specific shRNAs induce long-term (> 4 weeks) silencing at a cell:protocell ratio of 1:20. We are in the process of assessing the therapeutic potential of SC1-targeted, siRNA-loaded protocells using ex ovo avian embryos pre-infected with a NiV pseudovirus, which will enable us to optimize a protocell formulation for testing at BSL-4. Due to their enormous cargo capacity, as well as their stability and specificity, protocells show promise as delivery vehicles for therapeutic agents capable of preventing viral replication and transmission.

Although critical to the multifunctional materials of the future, the ability to integrate two disparate materials remains an enduring challenge. One approach to this problem is the use of combinatorial biology such as cell surface peptide display systems to screen a vast collection (or library) of unique peptides for those sequences with high binding affinity. These systems consist of large, diverse libraries (1-10 billion individuals) of peptides (7-15 amino acids) which are presented by a display scaffold protein hosted by a phage (virus), bacteria, or yeast cell. Presenting the peptides on a cell-based display scaffold allows for the peptides to be self-sustaining and aids in the identification of any specific peptides of interest. The peptide library can be rapidly screened for high affinity binders to a given target of interest, and those binders quickly identified through standard DNA sequencing techniques. Peptide display systems have traditionally been utilized in conjunction with organic-based targets, such as protein toxins or carbon nanotubes. However, this technology has been expanded for use with inorganic targets, such as metals, for biofabrication, hybrid material assembly and corrosion prevention. While most current peptide display systems employ viruses to host the display scaffold, we have recently shown that a bacterial host, Escherichia coli, displaying peptides in the ubiquitous, membrane protein scaffold, eCPX, can also provide specific peptide binders to protein target. We have extended the applications of the E. coli eCPX peptide library, for the first time, to an inorganic target. This peptide display system was used for the biodiscovery of aluminum binding 15-mer peptides. Through a process of biopanning involving four rounds of sorting, with increased washed stringency each round, aluminum specific peptide sequences were developed in less than one week&’s time. Monitoring of cells recovered from macroscopic aluminum targets after each round of sorting showed increased numbers of binders as the sorting process progressed. Furthermore, we also expressed phage-derived aluminum specific peptides on the E. coli eCPX scaffold and compared the binding to our peptide binders discovered using the E. coli eCPX system. We will detail the process of biopanning with macroscopic, inorganic targets, as well as binder isolation and discovery in a bacterial host. We will also present a computational approach which we used to explore the interaction of the discovered peptides with the aluminum surface.

The new paradigm referred to as e-Science, or data-intensive discovery, may bring unprecedented developments in science and technology with the huge amounts of data generated by digital instruments and sensor networks. For biosensors, in particular, data processing methods may be used to treat data from measurements with several principles of detection, including electrochemistry, electrical impedance and various spectroscopies. In this lecture, an overview will be provided of e-Science methods applied to biosensors made with nanostructured layer-by-layer (LbL) or Langmuir-Blodgett (LB) films, where synergy was sought in combining inorganic materials and biomolecules. Examples will be shown of multidimensional projection and parallel coordinate techniques, in addition to artificial intelligence methods, to enhance the performance of biosensors. The applications involved distinction between two tropical diseases in biosensors containing antigens immobilized in LbL films, elimination of cross talk in sensors based on field-effect devices and single molecule detection using surface-enhanced Raman scattering.

In this presentation we will describe the formation of fluid and resistive lipid membranes on nanoporous metal thin films. In our system, the nanoporous metal surface acts as the support as well as an electrode to characterize membrane resistivity. To achieve fluid and resistive membranes, a new two-step etching process to make nanoporous metal films with minimal cracking and pitting was developed. This two-step process can be generally applied as demonstrated by the formation of both nanoporous Au and Pt films. We utilize various lipid compositions to prepare lipid membranes that are both fluid and resistive, which mimics two keys properties of naturally occurring lipid membranes. Specifically, thiol-terminated lipids, DPPTE, are included in POPC vesicles that stabilize the lipid assemblies on the nanoporous gold surface and allow for lipid fluidity at room temperature. At the same time, DPPTE, which is in a gel-phase at room temperature, enhances the membrane rigidity that limits ion mobility across the membrane. SEM images were used to characterize the nanoporous substrate structure, while fluorescence recovery after photobleaching and electrochemical impedance spectroscopy were used to confirm lipid membrane fluidity and resistivity, respectively. The lipid assemblies supported on these types of nanoporous metal films described here provide a new platform for investigating lipid membrane properties, and potentially membrane proteins, which may lead to a variety of useful hybrid inorganic-biological architectures.

As a tool for massively parallel, high throughput molecular delivery, nanostructured platforms have emerged as a delivery technique with unique capabilities. Nanostructured platforms are able to access many different types of cells, even those that resist traditional techniques, and modern fabrication techniques are able to introduce unique functionalities into individual nanowires or nanotubes on the platform to enable electrical or optical access. Unfortunately the active delivery process, including the number or percentage of nanostructures that participate, the time scale of delivery, and the mechanisms, whether endocytotic or directly penetrating, are still unknown. Using single event, nanowire-based methods, these important characteristics of delivery are difficult to study and the technique is effectively a black box. We previously reported on a system, the nanostraw platform, capable of time resolved deliveries and in situ observation. Here we introduce a technique using nanostraws to directly observe molecular deliveries to cells. Using a simple, two part delivery assay, we demonstrate not only the direct observation of nanostraw penetration into cells, but also the probability and frequency of cell penetration by nanostraws. We are also able to demonstrate how soon cells accept nanostraw access after plating. Using these nanostraw techniques, we can make the first direct observations of the dynamic high-aspect ratio nanostructure to cell interface during molecular delivery.

Surface modifications of polydimethylsiloxane (PDMS) are fundamental for the exploitation of this versatile silicone in biological applications (E. Berthier et al., Lab Chip 12, 1224, 2012). Indeed, surface undergoes a typical phenomenon known as hydrophobic recovery, that poses severe time limitations to the observation of biological events and, in particular, to cell culturing. To date, countermeasures to hydrophobic recovery have been developed that allow cell culture for a period generally not exceeding one week (H. Ai et al., Cell Biochem Biophys 38, 103, 2003; C.C. Wu et al., Surf Coat Tech 205, 3182, 2011). A novel method of stable modification of PDMS surface chemistry was therefore elaborated, relying on the use of genipin, a natural cross-linker, and involving free amine moieties.
After plasma oxidation, surface silanization was performed to expose amine groups. Thereafter, collagen, a protein typical of the extracellular matrix, was covalently bound to thin PDMS films by exploiting genipin as a non-toxic cross-linking agent. Successful binding was verified with several independent tests, including water contact angle measurement, X-ray photoelectron spectroscopy (XPS), and fluorescence microscopy. Moreover, mechanical properties of the substrates were evaluated and found to be of the same order of magnitude of biological soft tissues (about 20 kPa). Collagen binding stability was observed for a period up to 30 days from PDMS modification. Protein presence on the surface was systematically investigated through fluorescence and XPS analysis, thus highlighting the efficiency of the proposed approach to hinder hydrophobic recovery of PDMS.
After assessment of chemo-physical properties, H9c2 muscle-like cells were cultured on modified membranes. The use of genipin as a cross-linker proved to be successful for a stable, non-citotoxic modification of PDMS surface chemistry. Modified PDMS films were indeed able to support long-term cell cultures and, in particular, to maintain the differentiated status of the selected cell line for a period of four weeks since differentiation induction.
On PDMS substrates, H9c2 myoblasts were able to fuse into long multi-nucleated myotubes, which were positive for myosin staining (a late muscle-cell differentiation marker). Analysis of confocal images allowed for a quantitative measure of actin and myosin colocalization to be performed, and this was found to be optimal for an efficient tissue functionality (G.G. Genchi et al., Coll Surf B, revision required).
New exciting perspectives for PDMS use can be therefore envisaged in a number of applications, ranging from muscle cell contractility studies to the development of bioMEMS and microfluidic devices (S. Sang and H. Witte, Microsyst Technol 16, 1001, 2010), the elaboration of drug screening protocols, and to bio-hybrid actuator design (J.C Nawroth, et al., Nat Biotechnol 30, 792, 2012).

The polymer/Pt tubular microrockets are synthesized using a template based electrodeposition method. The effects of different electropolymerized outer layers, including polypyrrole (PPy), poly(3,4-ethylenedioxythiophene) (PEDOT), polyaniline (PANI), poly(aminophenylboronic acid) (PAPBA) and of various inner metal surfaces (Ag, Pt, Au-catalase), upon the movement of such bilayer microtubes are evaluated and compared. Various electropolymerization conditions, such as the monomer concentration and medium (e.g. surfactant, electrolyte), have a profound effect upon the morphology and motion of the resulting microtubes. These oxygen bubble propelled Pt based microrockets are only 8 mu;m long, can operate in very low levels of the hydrogen peroxide fuel (down to 0.2%) and achieve a consistently remarkable high speed of 1400 body lengths/s in physiological temperature (the speed record of all artificial nanomotors). These microrockets can also move efficiently in various fuel enhanced biological media and can serve as an ideal platform for diverse biomedical and environmental applications. For example, lectin modified PANI/Pt microrockets are used for selective bacteria (E. Coli) isolation from food, clinical and environmental samples. Triggered release of the captured bacteria is also demonstrated by movement through a low-pH glycine-based dissociation solution. PAPBA/Ni/Pt microtube engines coupling the selective monosaccharide recognition of the boronic-acid-based outer polymeric layer with the catalytic function of the inner platinum layer. The resulting boronic-acid based microengine itself provides the target recognition without the need for additional external functionalization. 'On-the-fly' capture and transport of yeast cells (containing sugar on the cell wall) are illustrated. Release of the captured yeast cells is triggered via a competitive sugar binding involving addition of fructose. The SAM-coated Au/Ni/PEDOT/Pt microsubmarine displays continuous interaction with large oil droplets and is capable of loading and transporting multiple small oil droplets. These results demonstrate that such polymer based microrockets hold considerable promise for diverse applications, ranging from biomaterials isolation to environmental oil remediation.

Research at the nanoscale biotic-abiotic interface centers on endowing an inorganic nanocrystal with the physical, chemical, or energetic properties of biosystems. In this presentation we discuss the bioluminescence resonance energy transfer (BRET) between firefly luciferase from Photinus pyralis (Ppy) with core/shell semiconductive quantum rods (QRs). BRET has been studied as a function of QR aspect ratio and internal microstructure. The emission color of the QR was tailored from 600 nm to 800 nm by varying the QR morphology and composition. The QRs were found to be ideal energy acceptors, and Ppy-to-core distances were optimized by the synthetic control of rod morphology, surface chemistry, and Ppy:QR loading. The results were analyzed via Forster theory, lifetime measurements, and single-particle spectroscopy. The energy transfer efficiency, and BRET ratios, were found to be greater than the current state of the art. These results, and the potential applications will be discussed.

Recent developments in nanotechnology have brought us new advances in device fabrications and materials by emerging technologies from physicists, chemists, engineers, biologist, and materials science. There are a lot of challenges for material scientist to play an important role in this area since nanotechnology is a part of the chemical domain, which builds up materials at the molecular level. We present a hybrid approach of biological materials and biosensors based on molecularly imprinted polymer (MIP). MIP can be provided by “molecular imprinting technique,” which is a general protocol for the creation of “synthetic receptor or binding sites” with specific molecular recognitions in cross-linked network polymers. It has “high affinities binding sites” to detect specific biological molecules for fabricating advanced biosensors or chemical detection devices. Synthesis of “high affinity receptor sites” is a key contribute to achieve high sensitivity biological sensors based on MIP&’s molecular recognition functions. A hybrid approach of MIP&’s microfluidic synthesis was employed to synthesize high affinity binding sites. Hynrid microfabrication technology was also employed to offer us rapid assembly and integration of hybrid devices to fabricate MIP&’s based biological sensors, which satisfy a set of our multiple demands in nanotechnology.

Hybrid polymers based on polymethylsilsesquioxane (PMSSQ) are suitable as coating material enabling the variation of surface parameters and addition of new chemical functions. The inorganic PMSSQ is prepared by sol-gel process and mediates the binding to the surface and is responsible for the stability of the film. Organic side chains can be grafted onto functional side chains within the inorganic PMSSQ network. Controlled radical polymerization techniques enabled a covalent attachment of organic polymer chains. Utilizing the reversible addition-fragmentation chain transfer (RAFT) polymerization allowed the polymerization of reactive pentafluorophenyl acrylates (PFPA). The obtained poly(PFPA) can be converted with amines under mild conditions into a functional group after coating^1-3.
O-Nitrobenzyl (ONB) groups are light responsive and can be cleaved upon irradiation with ultraviolet light. Consequently, incorporation of ONB units between the inorganic and organic part enables the removal of the whole organic part.
The advantage using such hybrid systems is that a substrate independent coating process is available with only one precursor polymer, which is facilitated through the cross-linking of the PMSSQ. A curing step induces a secondary condensation and hence provides the mechanical stability. Afterwards, the coating can be functionalized by reaction with amines and exemplary stimuli-responsive films and target recognizing systems were prepared by this route. Thermo-responsive polymers with a reversible lower critical solution temperature phase transition such as poly(N-isopropylacrylamide) or pH responsive polymers based on chargeable poly(amines) have been investigated. The modification with target recognizing structures like biotin-streptavidin system enabled the immobilization of desired biological relevant compounds. In combination, these strategies were used to modify lab on chip systems. Combining these features with a light-sensitive group enhances the effect and allows removing functionalities afterwards by irradiating. Thus, target molecules can be immobilized as well as released by an external stimulus. Furthermore, the photocleavable group enables to influence structural wetting parameters by structuring techniques.
References:
1. D. Kessler, P. J. Roth and P. Theato: Reactive surface coatings based on polysilsesquioxanes: controlled functionalization for specific protein immobilization. Langmuir 25, 10068-10076 (2009).
2. D. Kessler and P. Theato: Reactive surface coatings based on polysilsesquioxanes: defined adjustment of surface wettability. Langmuir 25, 14200-14206 (2009).
3. D. Kessler, F. D. Jochum, J. Choi, K. Char and P. Theato: Reactive surface coatings based on polysilsesquioxanes: universal method toward light-responsive surfaces. Appl. Mater. Interfaces 3, 124-128 (2011)

Au nanostructures with highly branched surfaces have great potential for fabricating SERS-active tags for use in biomedical applications. The development of highly branched complex 3D nanostructures at a low cost, reduced eco-footprint, and well-defined hierarchical structure is fast emerging. In this study, a simple one-step approach to synthesize highly branched Au nanoflowers (Au NFs) has been proposed by using dopamine at room temperature. Dopamine is a monoamine neurotransmitter that plays an important physiological role in controlling the brain&’s reward and pleasure centers. The size and morphology of the Au nanoflowers could be easily controlled by tuning the concentration of gold precursors (HAuCl4) and dopamine by varying pH. Using scanning electron microscopy (SEM), the 3D flower-like particles were monodisperse (around 600-800nm in diameter), and had a large number of dendritic tips, 100-200 nm in length. The size distribution of the nanoflowers was characterized by dynamic light scattering (DLS). The average diameter was 850nm, consistent with the SEM analysis. The XRD pattern of the hierarchical Au nanoflowers showed sharp diffraction peaks exclusively attributed to Au crystals with the face-centered cubic (fcc) structure, which demonstrated the pure and well-crystallized Au. The formation of the Au nanoflowers was examined by conducting a time-lapse SEM study. Fourier transform infrared (FTIR) spectroscopy provided evidence that dopamine reduced Au ions to Au (0), and adsorbed on the Au nanoflower surface. In the typical SERS spectra of Au NFs and Au NPs (control) with adsorbed Rh6G, Au nanoflowers showed a significant enhancement of the SERS effect compared to gold nanoparticles. The cytotoxicity of Au nanoflowers was measured by the MTT assay in human lung cancer cells (A549), and mouse melanoma cell lines B16BL6 with various concentrations of Au nanoflowers. The results showed that the cell viability was more than 85%, demonstrating good biocompatibility. The intracellular uptake of the Au nanoflowers into A549 cells and B16BL6 cells was further investigated by dark field microscopy. We have demonstrated that the Au nanoflowers created in this study may be a promising candidate for Raman spectroscopy in living cells.

Biological organisms in flora and fauna have evolved a wide variety of micro- and nanostructures that provide unique optical signatures including outstanding, distinctive, dynamic coloration, high reflectivity or superior whiteness. Various intriguing photonic structures have been identified on the wings of insects, the torsi of spiders or the feathers of birds. More recently photonic structures have also been found in the shells or spines of marine animals. Life under water imposes very distinct constraints on organisms relying on visual communication and on the designs and the materials involved in aquatic photonic structures. Here, we present a bio-mineralized calcium carbonate-based crystalline photonic system buried in the shell of the blue-rayed limpet Ansates pellucida. The structure consists of layered stack of calcite lamellae with uniform thickness and inter-lamella spacing. This arrangement lies at the origin of the strong blue-green iridescence of the organism&’s characteristic stripes, which is caused by multilayer interference. Regular undulations of the multilayer in each stripe enhance the visibility in a wider angular range. This multilayer is supported by a disordered array of spherical particles with an average diameter of 300nm, most likely serving to enhance the contrast of the blue stripes. We present a full structural, elemental and optical characterization of this marine photonic system, supported by optical FDTD modeling.

Biocompatible, near-infrared luminescent gold nanoclusters (AuNCs) were synthesized in water using poly(ethylene glycol)-dithiolane ligands functionalized with carboxyl, amine, azide and methoxy groups. The AuNCs first underwent extensive characterization using UV-Vis absorption spectroscopy, steady-state luminescence and luminescence lifetime spectroscopy, along with high resolution transmission electron microscopy. The 1.5 nm AuNCs fluoresce at ~820 nm and have a quantum yield in the range of 4-8%, depending on the terminal functional group present while maintaining high colloidal stability (>1 year). The luminescence lifetime of the AuNCs exhibited a bi-exponential decay with time constants of ~ 1.4 µsec and ~ 0.2 µsec. The various terminal functional groups appended to the AuNCs can be easily conjugated to the biomolecules for biological applications without affecting the colloidal stability in various buffer conditions. We also characterized their two-photon absorption (TPA)-induced emission and measured the TPA cross-section of the AuNCs. The TPA cross section of AuNCs (Phi;σ> 140 GM @ 1000 nm) is much higher than that of organic dyes such as Rhodamine B. The large TPA cross section, combined with the significant luminescence quantum yield suggested that the AuNCs could be used effectively for cellular imaging. To evaluate this, AuNCs were injected directly into cells to confirm their long-term stability and high signal-to-noise ratio emission in the cellular environment. Next, the AuNCs were conjugated with cell-penetrating peptides (CPPs) and delivered to COS cells and imaged. The AuNCs exhibit minimal cytotoxicity due to the excellent biocompatibility imparted by the PEG ligands.

DNA is known primarily for its fundamental role in genetics, however, at a basic level, DNA is a polymer whose unique chemical recognition and organizational properties can be exploited for biological as well as non-biological purposes. The high specificity and programmability of DNA through base pairing, along with the nearly infinite range of sequence combinations, can be exploited in a variety of ways at the nanoscale to create and link organic and inorganic nanomaterials and nanostructures. In order to take full advantage of the potential of DNA assembly for nanodevice integration, the ability to precisely control the placement of DNA molecules and nanostructures on surfaces is crucial. In this work we describe an approach toward the controlled and ordered arrangement of DNA molecules, nanostructures and scaffolds on lithographically patterned, chemically (or biochemically) functionalized surfaces.
One approach that we have developed is based upon a “breadboard” consisting of nanometer-size metallic nanodots patterned by nanoimprint lithography and self-aligned pattern transfer. These nanodots serve as anchors for the assembly of 1D nanostructures (rigid DNA motifs and short carbon nanotube segments) through DNA hybridization or through a covalent bond. The dots are arranged in dimers, with the inter-dot spacing matching the length of the 1D nanostructures, which are functionalized with an active linker at their ends designed to bind to a complementary species on the nanodots. The nanodots are sufficiently small, ~ 5 nm, such that only a small number of 1D nanostructures can bind to them, allowing single-molecule control over the assembly.
In a second approach, we use electron beam lithography or nanoimprint lithography to pattern hydrophilic regions on a hydrophobic background. The regions are designed to match the size and shape of specific DNA nanostructures or DNA wrapped carbon nanotube segments which selectively bind to the hydrophilic areas. This approach is highly selective and can achieve nearly 100% yield with control over position and orientation.
By combining lithographic patterning with molecular recognition we are able to control the surface assembly of DNA nanostructures and DNA wrapped carbon nanotubes. This is an important step towards a broad range of applications, from DNA studies in the biomedical field to the use of DNA in developing future generation electronics.

Following biochemical reactions with high sensitivity and high spatial resolution is a required step to gain new insights in the identification and quantification of protein-DNA interaction mechanisms.
Here we propose a novel strategy based on Atomic Force Microscopy (AFM) to study at the molecular level the action of enzymes in confined monolayers of dsDNA substrates. We realized nanoarrays of vertically oriented, thiolated dsDNA molecules of variable density on an ultraflat gold film surface by means of AFM Nanografting and we investigated the mechanism of different enzymatic reactions (from restriction enzymes to helicases) on such nanostructures.
To follow the reaction of restriction enzymes, in particular DPNII and BAMHI, we registered with high precision the topographic height variation, before and after loading the enzyme, of engineered 44-mer DNA oligonucleotides, carrying the restriction sequence at the molecule half-height, as a function of DNA density in the patches. In the case of helicases we followed the unwinding kinetics of two homologous helicases belonging to the RecQ family, exploiting the different mechanical properties of double and single stranded DNA (the DNA flexibility reduces 50-fold from ds to ss).
We will show here that restriction reactions can be tuned in nanostructures by varying the DNA density. Experimental evidences and simple steric hindrance based models suggest that restriction enzymes exclusively gain access at the sides of laterally confined dsDNA monolayers and diffuse in a two dimensional fashion. Regarding RecQ helicases, we will show from a comparative assay that E.Coli RecQ activity is 10 times faster than the one of its human homologous RECQ1, Lack of C-terminus domain, different binding affinity and different cooperativity can explain the different processivity of the two helicases.
We will also provide preliminary results on the investigation of multiple reactions on the same DNA substrate, and on the parallel visualization of the action of enzymes on different DNA substrates (forks, blunt ended, etc.). Finally, we will discuss how to extend our work moving to real-time studies of enzymatic reactions on DNA functionalized gold nanoparticles.

There have been significant efforts in recent years in developing novel nanosystems for biomolecular engineering. We have designed near-IR fluorescent single-wall carbon nanotubes with molecular recognition domain by functionalizing them with nucleic acids. With molecular recognition and self-assembly capabilities of oligonucleotides, we have studied target-receptor interactions at the nanoscale. The thermodynamics and kinetics of these interactions can be extracted by utilizing the signal transduction from the optical nanomaterials. Our system is a powerful and unique optical platform that allows one to probe and analyze biomolecular reaction both in ensemble and at single molecule level.

Laser patterning of self-assembled monolayers represents a key step in many biofabrication schemes, e. g. in order to build up sensor arrays. The lateral resolution, however, usually is limited by optical diffraction. A means to enhance the lateral resolution takes advantage of nonlinear effects. Recent work in nonlinear laser patterning of self-assembled monolayers is presented. In photothermal processing a focused cw laser beam is used to locally heat the substrate surface and thermally initiate chemical reactions. For this reason, photothermal processing is highly nonlinear in laser power density and facilitates sub-wavelength patterning. Femtosecond laser processing, in turn, provides promising perspectives in sub-wavelength patterning of transparent via multiphoton absorption processes. Because of their exceptional stability, nonlinear laser processing of silane-based monolayers is most effective. In particular, photothermal laser processing at a wavelength of 532 nm and a 1/e spot diameter of 1.8 microns allows one to fabricate functional surface structures with lateral dimensions well below 100 nm. Such structures are used as templates to build up micro- and nanoarrays via local immobilization of proteins, DNA and other biomolecules.

The rapid synthesis of gold nanoparticles (GNPs) was done by solution plasma sputtering (SPS) process in the presence of a biopolymer, sodium alginate. We utilized the alginate polymer in order to both promote the generation of plasma in liquid environment and provide colloidal stability to the GNPs. The effect of sodium alginate concentration (varied as 0.2, 0.5, and 0.9 %w/v) and plasma sputtering time on the particle size of GNPs and the UV-visible absorption peaks were studied. The results indicated that preparation of GNPs in alginate gel matrix was succeeded by the SPS process in one step without any reducing agents. This technique is suitable for the production of GNPs with controllable size and high purity.

In recent years, a new generation of quantum confined colloidal semiconductor structures has emerged, with more complex shapes than simple quantum dots. These include nanorods and tetrapods. Beyond shape, it is also now possible to spatially vary the electron and hole potentials within these nanoparticles by varying the composition. Examples of these new structures include seeded dots, rods, and tetrapods, which contain a CdSe core embedded within a CdS shell. These structures may have many uses beyond those envisioned for simple quantum dots, which are frequently employed in luminescent applications. This talk is concerned with changes in the optoelectronic properties of tetrapods when the arms are bent. We demonstrate that seeded tetrapods can serve as an optical strain gauge, capable of measuring forces on the order of nanonewtons. We anticipate that a nanocrystal strain gauge with optical readout will be useful for applications ranging from sensitive optomechanical devices to biological force investigations.

Over the past several decades, supported lipid membranes have been used as model systems of cellular membranes, to investigate various membrane interactions, and as platforms for development of bio-sensors. Precise structural characterization by neutron and x-ray scattering offers a wealth of insight into membrane organization, self-assembly, and domain formation as well as how membranes respond to changes in their environment, e.g. temperature, protein binding, etc. In this talk, I will discuss some recent advances in our understanding of supported membranes including (1) high resolution details regarding the inorganic-organic interface, changes with in membrane structure with fabrication method, temperature, and solution conditions; (2) a novel, “textured” lipid phase induced by specific-multivalent protein binding to membrane embedded receptors, and (3) the potential use of polymer cushioned membranes in biotechnology applications. The talk will particularly highlight the importance of x-ray scattering techniques for structural characterization.

Artificial membrane systems allow researchers to study the structure and function of proteins in a matrix that approximates their natural environment, and to use these proteins in ex-vivo applications such as electronic biosensors, thin-film protein arrays, or bio-fuel cells. Most of these applications require control of protein orientation after immobilization onto device surfaces as the presence of two opposite protein orientations at best reduces the signal or at worst cancels it entirely. In this work, we have explored the role of the bilayer surface charge in determining transmembrane protein orientation and functionality during formation of proteoliposomes. We reconstituted a model ion pump, proteorhodopsin, in liposomes of opposite charges and varying charge densities and determined the resultant protein orientation. Antibody-binding assay and proteolysis of proteoliposomes showed physical evidence of preferential orientation, and functional assays provided evidence of vectorial ion transport in this system. Our results indicate that the manipulation of lipid composition can indeed control orientation of membrane proteins in liposomes. This technique opens up a simple route for controlling protein orientation in many applications ranging from solution vesicle assays to complex bioelectronic devices and sensors that use membrane proteins.

To improve patient survival rates against fatal diseases including cancer, advanced biomarker detection systems are needed for early-stage diagnosis, monitoring of therapeutic response, and determination of traeatment efficacy. For global applicability, advanced biosensor systems must also be easy to use, reliable, high-throughput, inexpensive to manufacture, and yet retain high sensitivity. Previously, we have demonstrated that chemically modified DNA-M13 bacteriophage can be used as a single bioanalytical platform that not only enables facile and rapid detection of antigens in solution but also provides modes for protein identification. After capturing the DNA-phage to magnetic beads by antigen binding, DNA conjugated gold nanoparticles were added, which upon hybridizing to the DNA-phage led to an immediate visible decrease in color in the solution. These DNA gold nanoparticles could also be reversibly released from the phage and then captured to a DNA microarray for antigen typing. To further increase sensitivity and still allow for multiplexed detection, we demonstrate here our recent efforts on using DNA modified bacteriophage and DNA conjugated SERS (Surface Enhanced Raman Spectroscopy) active Au/Ag core shell nanoparticles. We modified the major capsid proteins of M13 bacteriophage with DNA using amine-succinimidyl ester and hydrazone-aldehyde chemistry. The binding capacity of modified DNA phage to DNA-SERS active nanoparticles were determined by negative stained TEM images. For the sensing assay, the model antigen biotinylated-anti-goat IgG was captured onto streptavidin coated 5 micron silicon pillars. Capturing antigen, DNA-phage and subsequently the DNA-conjugated SERS particles to 5 micron silicon patterns allowed for easy analysis by Raman spectroscopy. The DNA conjugated M13 phage were next reacted with the captured antigen, followed by addition of the DNA conjugated SERS active nanoparticles and Raman spectroscopy measurements. By measuring SERS intensities as a function of antigen in solution, number of DNA strands per phage, and maximal loading of the SERS active particles per phage, antigen amounts could be quantitatively compared to the standard curve obtained of SERS intensity versus number of SERS nanoparticles. Based on our measurements, our bacteriophage biosensor system was able to detect femtomolar amounts of proteins.

Hydrogen sulfide (H2S), a highly toxic gas, is a by-product of the petroleum, paper, mining, and water treatment industries. Exposure to low levels of H2S (< 5 ppm) is innocuous; however exposure to higher levels of H2S may cause irritation to eyes and lungs and, in the extreme, result in respiratory arrest and death. Sensitive H2S sensors, able to continuously monitor gas levels, are a necessity for workers in these industrial environments. Nanostructured gold is a promising candidate for low power, sensitive H2S sensors. The high chemical affinity between gold and sulfur enables adsorption of H2S thereby increasing the grain-to-grain barrier to electron transport and, ultimately, the electrical resistance. Unlike other commonly used gas sensor materials which must be heated during sensing operation, gold can be used at ambient temperature which significantly reduces the power consumption of the sensor. Furthermore, the high surface-to-volume ratio of nanoscale gold increases the volumetric number of H2S reactive sites enhancing sensitivity over its bulk counterparts. Bio-templated assembly is a technique that allows facile bottom-up assembly of nanostructures at ambient temperature and mild conditions. Specifically, the M13 filamentous bacteriophage, approximately 880 nm in length and 6 nm in diameter, has been used to assemble various inorganic nanostructures through the display of material specific binding peptides on its viral protein coat. In this report, we have fabricated a H2S gas sensor using M13-templated gold nanowires. A genetically-modified, gold-binding M13 template was deposited on an insulating substrate between two pre-patterned gold electrodes. Chains of gold nanoparticles were assembled along the length of the template. Gold electroless deposition was used to increase and control the connectivity of the nanoparticles, tuning the electrical conduction of the resulting nanowires for optimum sensing performance. Scanning electron microscopy was used to characterize the morphology and connectivity of these nanowires across the electrodes. The electrical resistance of the viral-templated gas sensors was measured as a function of temperature to determine the dominant conduction mechanism. The resistance change, response time, recovery time, and sensitivity to H2S gas were measured from 25 ppb to 40 ppm at room temperature while applying a constant bias. The M13 bacteriophage template enabled facile, controlled assembly of gold nanocrystals for the realization of sensitive H2S sensors.

Immobilization of proteins in porous materials has attracted attention of many
researchers due to the beneficial properties of the pore system and its interaction
with the proteins. The confinement effect in porous materials, whose pore sizes
match the size of many enzymes, usually stabilizes the enzyme but leads to
difficulties in diffusion as well as to lower loadings. Macroporous materials can
overcome these limitations but that results in lower protein stabilization and
leaching.1,2
We chose different porous materials to optimize the immobilization and performance
of Glucose-6-phosphate dehydrogenase (G6PDH). In synthetic enzymatic pathways
it is one major enzyme that regenerates NADPH. It is a very large 104 kDa dimeric
protein with a hydrodynamic radius of approximately 13 nm.
Mesoporous siliceous cellular foams (MCF) and hierarchical meso- macroporous
carbons are tuned to efficiently immobilize G6PDH. These materials offer a variety of
functional groups and surface characteristics. Titrations of the zeta potentials of each
material are used to investigate the surface charges and to determine the isoelectric
point of each porous support. The information allows matching the charges of
G6PDH (pI = 4.6) and the supports during the immobilization process.
The uptake of the enzyme is monitored for each material. The uptake curves give a
first clue which interaction may apply and how strong they are. The materials that are
negatively charged at the pH of immobilization exhibit low uptake due to the repulsion
with the negatively charged protein. Positively charged silica materials show
according to their zeta potential the highest capacities for G6PDH immobilization at
pH 7.4. A large fraction of the protein is attached to aminopropyl-modified MCF
(MCF-NH2) by adsorption. The electrostatic interaction is very strong and no leaching
was detected. The activity was almost completely retained. The covalent
immobilization by Schiff-base reaction to a glutaraldehyde-modified support
(MCF-glu) was very fast. The uptake by hierarchical porous carbons is low but it
increases after oxidation. This indicates that the enzyme is not attracted by
hydrophobic forces to the carbon materials.
The prepared biohybrid materials exhibit excellent stabilities as well as enzyme
uptake properties. The efficient generation of NADPH was performed with the
immobilized G6PDH.
(1) S. Hudson, J. Cooney, E. Magner Angew. Chem. Int. Ed. 2008, 47, 8582-94.
(2) C.-H. Lee,T.-S. Lin, C.-Y. Mou Nano Today. 2009, 4, 165-179.
(3) D. I. Fried, K. Tropp, M. Fröba ChemCatChem, 2012, in press.
Acknowledgments: We thank the Landesexcellence Initiative Hamburg (LEXI) for financial support.

Self-assembled monolayers (SAMs) are an inexpensive and versatile way of introducing reactive groups to a surface that are capable of covalent attachment of functional bio-molecules.[1, 2] In immunosensors, antibodies are used as the recognition element of the sensor through binding to a specific target antigen. Antibody arrays enable parallel detection of multiple proteins in low sample volumes and can be used to identify disease biomarkers valuable for diagnosis, one such application is in cancer proteomics.[3] Current immunosensor research is focused towards the development of point of care devices.[4]
In this contribution we report the use of a photo-labile SAM to selectively pattern antibody arrays. The SAM can be photo-deprotected using near-UV light to expose reactive surface groups that can covalently bind protein streptavidin (STV) and human recombinant single chain fragment antibodies (scFv) on an oxide substrate. This covalent attachment leads to photo-patterned arrays of surface bound antibody fragments that have been used for specific detection of disease biomarker proteins in buffer and in human serum. The use of blocking reagents to reduce non-specific adsorption of proteins to patterned substrates will also be discussed.
1. C. D. Hodneland, Y. S. Lee, D. H. Min, M. Mrksich, Proc Natl Acad Sci U S A 2002, 99, 5048.
2. G. B. Sigal, C. Bamdad, A. Barberis, J. Strominger, G. M. Whitesides, Anal Chem 1996, 68, 490.
3. E. Kopf, D. Zharhary, Int J Biochem Cell Biol 2007, 39, 1305.
4. T. R. Holford, F. Davis, S. P. Higson, Biosens Bioelectron 2012, 34, 12.

Controlling the electrical contact of redox enzymes with electrodes is a critical issue for enzymatic biodevices such as biofuel cells and biosensors. The mutual positioning between enzyme molecules, mediator molecules (not always necessary), and electrode surface determines the efficiency, reproducibility, and stability of the bioelectrocatalysis systems. We present herein an enzyme/mediator/electrode ordered ensemble that shows both “high turnover rate” and “large catalytic current”. In order to satisfy both of these requirements, the larger amount of enzymes than monolayer should be immobilized with keeping effective contact with electrodes. We realize such ideal condition by taking advantage of a film of well-aligned carbon nanotube forest (CNTF) [1] consisting of single-walled CNTs arrayed with a pitch of 16 nm. We have previously reported the incorporation of fructose dehydrogenase (FDH) and Laccase (LAC) as the anodic and cathodic electrode catalysts, respectively. [2] By connecting the FDH-CNTF anode and the LAC-CNTF cathode showed the world highest power density of 1.8 mW cm-2 (at 0.45 V) in stirred 200 mM fructose, 84% of which could be maintained after continuous operation for 24 h.
In this work, the molecularly ordered composites of polyvinylimidazole-[Os(bipyridine)2Cl] (PVI-[Os(bpy)2Cl]) and glucose oxidase (GOD) are assembled inside a film of CNTF. The structure of the prepared GOD/PVI-[Os(bpy)2Cl]/CNT composite film is entirely uniform and stable; more than 90% bioelectrocatalytic activity could be maintained even after storage for 6 days. [3] Owing to the ideal positional relationship achieved between enzyme, mediator and electrode, the prepared film shows a high bioelectrocatalytic activity for glucose oxidation (ca. 15 mA cm-2 at 25 °C) with an extremely high electron turnover rate (ca. 650 s-1) comparable to the value for the GOD solutions, indicating almost every enzyme molecules entrapped within the ensemble (ca. 3 × 1012 enzymes in a 1× 1 mm2 film) can work to the full. The free-standing, flexible composite film can be used by winding on a needle device, for example, a self-powered sugar monitor, that changes the blinking interval of an LED upon simply being inserted into a grape.
[1] D. N. Futaba, K. Hata, T. Yamada, T. Hiraoka, Y. Hayamizu, Y. Kakudate, O. Tanaike, H. Hatori, M. Yumura, S. Iijima, Nat. Mater. 2006, 5, 987.
[2] T. Miyake, S. Yoshino, T. Yamada, K. Hata, M. Nishizawa, J. Am. Chem. Soc., 2011, 133, 5129.
[3] S. Yoshino, T. Miyake, T. Yamada, K. Hata, M. Nishizawa, Adv. Energy Mater., DOI: 10.1002/aenm.201200422.

Nowadays, a hybrid device approach involving the integration of biomolecules and nano-objects (e.g., nanoparticles, nanotubes) with macroscopic electronic transducers is considered as one of the most attractive platforms for the transformation of (bio-)chemical information into usable electrical signals. In this context, the coupling of nanoparticle/biomolecule inorganic/organic hybrids with semiconductor field-effect devices based on an electrolyte-insulator-semiconductor (EIS) system could open new and very promising opportunities for biosensors and biochips.
In this work, we present a new capacitive EIS sensor functionalized with ligand-capped gold nanoparticles (AuNP) capable for a label-free electrostatic detection of charged macromolecules by their intrinsic molecular charge. The EIS sensor detects the charge changes in the AuNP/macromolecule inorganic/organic nano-hybrids induced by the adsorption or binding events. Since the surface of AuNPs can be easily modified with various charged ligands of interest, the use of AuNPs offers many advantages in terms of recognition and transduction of molecular binding events. Moreover, because the vast majority of biomolecules are charged in physiological conditions, the AuNP-modified field-effect devices can provide an universal and efficient platform for detecting a wide variety of charged biomolecules.
The generic approach has been exemplary demonstrated by realizing capacitive EIS sensors for the electrostatic detection of cytochrome c, DNA hybridization, poly-D-lysine as well as the layer-by-layer adsorption of polyelectrolytes. For the experiments, the surface of an EIS sensor consisting of Al-p-Si-SiO2 structure has been silanized and modified with citrate-capped AuNPs (15-20 nm). The surface of the functionalized EIS sensor has been characterized by scanning-electron microscopy and atomic-force microscopy method. The sensors were characterized in buffer solutions by means of capacitance-voltage and constant-capacitance methods before and after the adsorption or binding of charged macromolecules on the surface of AuNPs. For comparison, the EIS sensors without AuNPs have been tested, too. An electrostatic model for a capacitive field-effect EIS sensor functionalized with AuNP/charged-macromolecule hybrids will be presented.

Photosystem I (PSI) is a protein complex found in the thylakoid membrane in plants that is responsible for electron excitation in the process of photosynthesis. The extraordinary internal quantum efficiency of PSI, approaching 100%, makes PSI a unique material to study in non-biological settings. A collaboration between the Cliffel and Jennings research groups here at Vanderbilt University has developed a facile method for depositing thick films of this photoactive protein on the surface of electrodes. We recently published work in which a platinum catalyst could be integrated with a film of PSI on a gold electrode for solar energy conversion. Here we will describe how tremendous photocurrent enhancements are possible by using a semiconducting electrode rather than metal electrodes. Furthermore, we demonstrate how this interface may be investigated by changing the doping type and density of the semiconductor as well as the electrochemical mediator.

Cell-based tissue engineering strategies may address the limited availability of bone graft material required to heal large bone defects. However, success in small animal models is variable and the process of regeneration in scaffold materials is not yet well understood. Here we describe the biomimetic rationale, fabrication, and in vivo performance of a collagen-hydroxyapatite scaffold for bone regeneration.
Bone is primarily composed of type I collagen mineralized with carbonated apatite. To mimic the physiological composition of bone, we have developed a collagen-hydroxyapatite scaffold and demonstrated healing of a critical size defect in the mouse calvarium. This biomimetic platform is relatively simple to fabricate, and is both highly osteogenic and osteoconductive. Furthermore, cells can easily remodel the collagenous phase through secretion of matrix digesting factors.
In addition to examining the effect of cell source on bone tissue engineering utilizing a collagen-hydroxyapatite scaffold, we also wondered about the progression of regeneration within such a material. Towards this end, we have established an imaging platform to view cells within a scaffold construct at multiple timepoints in vivo. This model has provided an unprecedented view of the progression of bone repair within a tissue-engineered construct. Using transgenic reporter mice to distinguish resident and scaffold-seeded bone forming cells, we show for the first time, the dynamic interplay between these two populations over the course of weeks.