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Although it is well recognized that cell-matrix interactions are based on both molecular and geometrical characteristics, the relationship between specific cell types and the three-dimensional morphology of the surface to which they are attached is poorly understood. This is particularly true for glomerular podocytes - the gatekeepers of glomerular filtration - which completely enwrap the glomerular basement membrane with their primary and secondary ramifications. Nanotechnologies produce biocompatible materials which offer the possibility to build substrates which differ only by topology in order to mimic the spatial organization of diverse basement membranes. With this in mind, we produced and utilized rough and porous surfaces obtained from silicon to analyze the behavior of two diverse ramified cells: glomerular podocytes and a neuronal cell line used as a control. Proper differentiation and development of ramifications of both cell types was largely influenced by topographical characteristics. Confirming previous data, the neuronal cell line acquired features of maturation on rough nanosurfaces. In contrast, podocytes developed and matured preferentially on nanoporous surfaces provided with grooves, as shown by the organization of the actin cytoskeleton stress fibers and the proper development of vinculin-positive focal adhesions. On the basis of these findings, we suggest that in vitro studies regarding podocyte attachment to the glomerular basement membrane should take into account the geometrical properties of the surface on which the tests are conducted because physiological cellular activity depends on the three-dimensional microenvironment.

Here we present a new bonding protocol for SU-8 negative tone photoresist that exploits the chemical modifications induced in the resin by exposure to 254 nm (UVC) light. Fourier Transform Infrared microspectroscopy (mu-FTIR) was used to carry out a thorough study on the chemical processes and modifications occurring within the epoxy resin by exposure to 365 nm and 254 nm light. In particular, we established that UVC light promotes the opening of the epoxy rings bypassing the post-exposure bake. The possibility to promote a further activation of the resin, already patterned with standard UV lithography, was exploited to produce closed microfluidic devices. Specifically, we were able to fabricate fluidic chips, characterized by broadband transparency from mid-IR to UV and long term stability in continuous flow conditions. CaF2 was used as substrate, coated by sputtering with a nanometric silicon film, in order to make surface properties of this material more suitable for standard fabrication processes with respect to the original substrate. The fabricated microfluidic chips were used to study by mu-FTIR the biochemical response of live breast cancer MCF-7 cells to osmotic stress and their subsequent lysis induced by the injection of deionized water in the device. mu-FTIR analyses detected fast changes in protein, lipid and nucleic acid content as well as cytosol acidification.

In this contribution we present the first example of a microfiuidic chip based on BaF2 for Infrared Microspectroscopy (IBMS) of living cells. The advantage in using barium fluoride as platform relies on its high IR transparency, especially in the spectral region below 1300 cm(-1), where the absorption bands of nucleic acids and carbohydrates are located. Barium fluoride is slightly soluble in water (0.12 g/100 g water) and it is potentially harmful for living cells. To overcome these problems, here we exploit an approach whose feasibility has been demonstrated previously on CaF2: the surface modification obtained by sputtering a thin Si layer on the surface. The Surface Modified Microfluidic Devices (SM-MD) hence obtained not only solve the BaF2 drawbacks, but also provide a silicon-like substrate fully compatible with standard micro-fabrication processes. These potentialities are here further explored in the direction of chemical or topographical nano-patterning of the silicon-like surface. The silicon thin layer was structured in the shape of 300 nm wide grooves (500 nm pitch) with a thickness of 35 nm by using standard NIL and etching processes; chemical patterning was achieved by exploiting silane chemistry. Finally, we tested the performances of these devices at SISSI beamline@Elettra, collecting IR spectra of single MDA-MB-231 living cells maintained either in physiological solution or complete medium. A comparison of the spectra of a single cell obtained in BaF2 and CaF2 MDs is reported. (C) 2013 Elsevier B.V. All rights reserved.

Presently, no resist material has suitable properties to satisfy simultaneously all requirements on lateral resolution and plasma etching resistance for pattern transfer of high aspect ratio nano-structures. Herein, a recent innovative alumina spin-on material for use as both positive and negative high resolution resist for electron beam lithography (EBL) is presented. This system turns into an almost completely inorganic alumina system when exposed to the electron beam, showing exceptional selectivity (100:1), compared to even the most robust polymeric resist, when used as mask for dry etching of silicon in fluorinated plasmas, obviating the need for a hardmask layer deposition. A full characterization of the resist selective removal under different processing conditions, such as electrons acceleration voltage and post-exposure bake temperature and time, is presented. Owing to the high selectivity and high resolution, silicon nano-structures of size down to 11 nm and high aspect ratio (up to 20) have been obtained for the first time in a one-step process.

We report the nanopatterning of the methylammonium lead iodide (MAPbI3) perovskite, a polycrystalline hybrid organic-inorganic semiconductor with very promising perspectives for photovoltaic and optoelectronic applications. Nanopatterning of MAI is obtained via Pulsed-NIL technology, an ultrafast version of the thermal nanoimprint lithography (NIL) based on stamps with integrated heaters. By Pulsed-NIL we were able to replicate onto the hybrid perovskite structures with details at the sub-100nm scale, in spite of its crystalline nature. This work shows a new possibility for the exploitation of organo-metal halide perovskites in optoelectronic devices with more complex architectures than just planar films. Display Omitted Methylammonium lead iodide thin film nanoimprinting approach is proposed.V-groove nanostructures are transferred to the photoactive material via Pulsed-NIL.The technique allows to perform the step in less than 1s.The preliminary trials show the possibility to scale up the process.

Catalyst nanoparticles are required in most established synthetic routes for the growth of carbon nano-tubes (CNTs) and semiconductor nanowires (NWs). A simple and widespread method to coat a substrate with metal nanoparticles is based on the dewetting of a thin metal film, often induced by a process of thermal annealing, and resulting in the formation of fine droplets. However, the control of the sole thermodynamic parameters affecting the dewetting results in a rather broad size distribution and random positioning of the catalyst. Efforts aiming at a better control of size and position of the catalytic seeds by top-down and bottom-up patterning techniques are well documented. However, patterning the catalyst may be a useless process step if, once approaching the temperatures at which the growth of nanowires and CNTs is catalyzed, nanoparticles become mobile on the substrates, displacing from their position and altering the initial size distribution due to coalescence. Moving from this observation, we propose a simple and general method to prevent the diffusion of the catalyst during annealing steps or the initial stage of the synthesis of NWs and CNTs. Its principle consists of confining the catalyst nanoparticles on top of vertical columnar structures. This can be achieved by etching the substrate all around the nanoparticles by dry anisotropic etching using the nanoparticles themselves as etch masks. Nanopartides thus sitting on top of nanopillars result effectively confined, even at temperature at which they are found highly mobile on flat substrates of the same material, due to surface tension effects. Experimentally, we demonstrate this principle with iron nanodots obtained on silicon by lift-off starting from a P(MMA-b-S) diblock-copolymer template. The iron nanodots isolated on top of columnar nanostructures, were found to be stable at 600 degrees C, temperature suitable for the growth of CNTs. (C) 2013 Elsevier B.V. All rights reserved.

Nanoelectrode arrays (NEAs) are increasingly applied for a variety of electroanalytical applications; however, very few studies dealt with the use of NEAs as an electrochemical generator of electrogenerated chemiluminescence (ECL). In the present study, arrays of nanodisc and nanoband electrodes with different dimensions and inter-electrode distances were fabricated by e-beam lithography on a polycarbonate layer deposited on boron-doped diamond (BDD) substrates. In particular, NEAs with 16 different geometries were fabricated on the same BDD sample substrate obtaining a multiple nanoelectrode and ultramicroelectrode array platform (MNEAP). After electrochemical and morphological characterization, the MNEAP was used to capture simultaneously with a single image the characteristic behaviour of ECL emission from all the 16 arrays. Experiments were performed using Ru(bpy)(3) (2+) as the ECL luminophore and tri-n-propylamine (TPrA) as the co-reactant. With a relatively limited number of experiments, such an imaging procedure allowed to study the role that geometrical and mechanistic parameters play on ECL generation at NEAs. In particular, at high concentrations of TPrA, well-separated individual ECL spots or bands revealed an ECL signal which forms a pattern matching the nanofabricated structure. The analysis of the imaging data indicated that the thickness of the ECL-emitting zone at each nanoelectrode scales inversely with the co-reactant concentration, while significantly stronger ECL signals were detected for NEAs operating under overlap conditions. Supplementary material: [http://cherry.chem.bg.ac.rs/handle/123456789/3566]

We have successfully grown regular arrays of InAs/GaAs quantum dots on patterned Si substrates. Thanks to the capability of nanoimprint lithography, we were able to obtain uniform patterns extended over some cm(2) areas, with periods of 300 nm. Ex-situ and in-situ treatments of the surface allowed us to completely remove any residual oxides prior to growth without the use of hydrogen beams, at temperatures compatible with standard III-V molecular beam epitaxy. The growth protocol was optimized in order to obtain a perfect selectivity of InAs/GaAs nanostructures in the holes, without any deposition on the planar areas. (C) 2015 Elsevier B.V. All rights reserved.

Natively porous hybrid organic-inorganic sol-gel systems have been engineered to be used as functional positive photoresists, aimed to the realization of microsensors in a single-step process. Interesting results have been obtained combing three main sol-gel system features: the direct pattern-ability through X-ray lithography, being processable without the addition of a photocatalyst; the functionalizability, properly designing the organic component of the hybrid network or incorporating active species; the open micro or mesoporosity, tailored by the synthesis process and precursor choice. The photoprocessable porous films have been synthesized starting from a Bridged Polysilsesquioxane (BPS) precursor, 1,4-bis(triethoxysilyl)benzene. The correlation between chemical properties of the solgel material and its patternability is described in detail. X-ray exposure leads to a progressive alkyl and aromatic compound elimination and promotes inorganic condensation in the system, allowing the selective dissolution of irradiated cross-linked films in suitable etchants. Patterns of final resolution lower than 100 nm have been realized on BPS-based films synthesized in acid conditions, a procedure that allows to take advantage of a straightforward embedding protocol for active species in the sol-gel matrix. The BPS-based system has been doped with a covalently linked quinolinium dye, obtaining thin sensing films patternable by X-ray lithography. A feasibility test for the fabrication of optical microdevices, where fluorescence properties are obtained directly on the patterned coatings, has been provided. (C) 2010 Elsevier B.V. All rights reserved.

Hybrid sol-gel materials are suitable dielectric materials for a range of optical applications, due to the versatility by which their optical and mechanical properties can be adjusted and the easy patterning by various lithographic techniques. We report on the synthesis of engineered epoxy based hybrid organic-inorganic sol-gel materials and on their lithographic processing, specifically applied to the fabrication of microlens arrays. The investigated sol-gel systems exhibit enhanced transparency in the near UV region with respect to most commercial thermoplastic polymers and sol-gel materials, used in embossing and nanoimprinting technology; in addition, they offer a better shape stability and superior mechanical properties, such as higher elastic modulus and hardness, improving the perspectives of the introduction of microstructured and nanostructured surfaces in applications where resistance to environmental mechanical-chemical degradation mechanisms is highly desirable, in particular for outdoor applications.

Engineering of the cellular microenvironment has become an attractive strategy to guide cellular activities such as spreading, motility, proliferation and differentiation. From a technological perspective, the physical crosstalk between a cell and its surrounding represents a design parameter that may be modulated to achieve desired osseointegration in orthopedic and dental implants. In this study we use a surface engineering approach to tap into the interaction between the cell and its surroundings in order to modulate osteogenic adhesion-dependent differentiation and bone tissue formation. The effectiveness of this approach was studied by observing the in vitro cellular behavior of human osteoblastic cells (SaOs-2) seeded on silicon substrates with different nano-scale surface patterns. Our findings suggest that substrate nanopattern geometry can differentially control early cell differentiation decisions. Interestingly, rescue of the differentiation process by supplementation of growth medium with soluble differentiation factors did not appear to compensate for the differences in the early cellular fate decisions observed on the different patterns. Critically, combining surface topography and soluble differentiation factors appeared to modulate the speed of the differentiation process and its shut down at the end of terminal differentiation. Taken together, our findings suggest that cells recognize physical nano-scale topography as an instructive signal that is integrated to fine-tune the multimodal control and guidance of cell behavior and cell fate. This information may find utility in the design of orthopedic and dental implants or innovative bio-materials for regenerative medicine.

A relevant increase in the electroanalytical application of sub-micrometer sized electrodes came with the development of arrays of nanoelectrodes. In the nanoelectrode ensembles (NEEs), the spatial distribution of the nanoelectrodes is random, while in the nanoelectrode arrays (NEAs) it is ordered. NEAs are typically fabricated by top-down microfabrication techniques, such as electron-beam lithography or nano-imprinting, while bottom-up technologies, such as the template methods and self-assembly are more often used for NEEs. NEEs/NEAs can exhibit three distinct response regimes depending on the scan rate or distance between the nanoelectrode elements: Total Overlap, Pure Radial and Linear Active. In the present work, NEAs are prepared by e-beam lithography and used for voltammetric measurements.

Organic photovoltaic cells (OPV) are among the most promising systems for energy extraction and conversion from renewable energy sources. However, major problem to be solved before industrial production could become economically viable is represented by their still low conversion efficiency. The organic solar cell architectures are presently the result of a compromise between achieving complete light absorption using active layers that are thicker than the optical absorption length and achieving efficient charge collection at the electrodes which is favoured in thinner layers. We present a concept and its experimental demonstration that would solve efficiently the above trade-off problem by making use of a new type of light trapping elements. The simple fabrication scheme, based on a self-aligned UV exposure process, suggests its potential up-scalability to large systems, at low production cost.

We have developed a fabrication methodology that relies on nanoimprint lithography (NIL), wet etching (Buffered Oxide Etch solution) and plasma etching in an Inductively Coupled Plasma (ICP); with our process we are able to pattern extended surface areas of about 1cm^2 with an array of rectangular channels or hemispherical holes of nanometric size. The fabricated samples are used in adsorption experiments to test recent theoretical predictions concerning the capillary condensation in capped capillaries.

We present the first results of microstructure replica with re-usable masters in poly vinyl alcohol (PVA). Microneedles of heights ranging from 500 to 1100@mm were fabricated through double-exposure deep X-ray lithography (DXRL) process and successfully replicated. A single PVA master was used as a template to successfully replicate up to 10 PMMA microneedle arrays, suggesting that this method might be a possible alternative to PDMS processes. A preliminary characterization of the relationship between the surface roughness of the substrates and the force required for demoulding was also performed.

In this paper, we present a novel approach toward the fabrication of mechanical oscillators with a local chemical functionalization with subnanometric control. Our method exploits the reactivity of the freshly cleaved surfaces that form when a monocrystalline silicon microstructure is cleaved. The surfaces that form after the cleave expose an atomically smooth Si(111) surface which, if a suitable chemical environment is provided, could undergo to a cycloaddition process that create stable, local and specific chemical bonds. Here, we demonstrated the feasibility of such a scheme on a twin cantilevers geometry, we prove the effective selectivity and stability of the cycloaddition process and we provide experimental evidence that below a critical size, the cleavage procedure creates step free atomically flat silicon surfaces.

Islet transplantation has great potential in restoring normoglycemia in diabetic patients. However, current clinical islet transplantation is associated with high graft failure the reasons of which still remain obscure. It is noteworthy that the process of islet isolation exposes the islet to a variety of cellular stresses, including disruption of the cell-matrix relationship, an event associated with induction of apoptotic pathways. This may adversely affect islet viability and function leading to the observed failure of islet transplantation. The cell-matrix relationship, characterised by adhesion interactions between membrane integrin receptors and the extracellular matrix molecules at the nanoscale, is known to modulate cell survival and growth. The aim of this study is to present a unique method of fabricating large area artificial substrates with biomimetic nanoscale surface topography. The substrates were realised by thermal evaporation of tin on silicon wafers to form randomly assembled nano-islands. Using the nano-islands as masks, the wafers were plasma etched to transfer the pattern onto silicon. We then examined the hypothesis of whether beta-cell function and survival can be enhanced by optimized restoration of cell-matrix adhesion. The results of in vitro cell cultures show that insulinoma cells significantly adhered and enhanced insulin secretion when cultured on these nano-structured substrates compared to planar controls. With this unique fabrication process we propose a simple, robust and accessible technology for fabricating platforms for potential use in the investigation and modulation of early pancreatic beta-cell behaviour which may find utility in the development of strategies for pancreatic islet engineering and transplantation.

A major limitation in the translation of stem cells technology to clinical applications is the lack of efficient control over their proliferation and differentiation. Recent in vitro research findings suggest that biomaterials with nanoscale surface topography can influence cell behaviour like adhesion, proliferation and differentiation. Therefore, the identification of biomaterials that support appropriate ES cell attachment, proliferation and differentiation into cells of interest is an attractive strategy worth investigation. In this study we present the design and fabrication of thin films of gold with surface topography of varying roughness using a combination of microfabrication techniques. We then explored their biomimetic potential to direct differentiation of ES cell-derived neural precursors. Standard glass coverslips and tissue culture plastic were used as reference materials. Our preliminary results show that ES-derived neural precursors best adhered on gold and underwent the highest differentiation on gold films with root mean square surface roughness (Rq) of 21 nm (72 ± 6%) after five days of culture in the absence of traditional soluble neurotrophic factors. Moreover, when cells were seeded on a combination of micro-scale grooves with nanoscale surface roughness, axonal outgrowth orientation was observed to be influenced by the grating axis. This data identifies gold as a potential biomaterial candidate with suitable cytocompatibility for ES cell research. It further lends support to the hypothesis that biomaterial design may optimize ES cell-derived neural differentiation and organisation. This may find utility as a tool to synergically complement soluble chemical factors currently used in in vitro protocols for the stimulation and modelling of ES differentiation. Ultimately, substrate patterning may hold special utility in the design of neural prostheses because repair of neurological injuries requires directional guidance.

The fabrication of single electron transistors and/or highly sensitive biosensors is still a challenging task on account of the tight control required to get proper shapes and size of the electrodes. The nanosized tips and the separation of a few nanometers between electrode pairs are critical features. Conventional lithography is not suited to obtain these features because of the resolution limits, so that previous alternative approaches have involved the use of electron beam lithography, focused ion beam lithography or scanning probe nanolithography. The novel approach presented in this letter is the exploitation of X-ray lithography in the Elettra synchrotron to fabricate arrays of nanocontacts spaced a few nanometers, devoted to the design of a new class of nanodevices based on nanoparticles and/or single molecules, including single electron transistors and highly sensitive biosensors. The method to fabricate such devices is illustrated and discussed. Experimental details of the fabrication process are given and preliminary results are presented through SEM and AFM images. It is worth noting that this paper presents a viable method to produce nanocontacts by using the X-ray lithography by synchrotron radiation source, that has not yet been reported together with experimental, though preliminary, data. (c) 2006 Elsevier B.V. All rights reserved.

Microcontact printing (μCP) is very attractive as a low cost, high throughput, and sub-100-nm resolution lithography. However, macroscopic distortions (thermal expansion, swelling by the solvent-assisted procedure of inking, externally induced strain) as well as microscopic distortions (membrane relaxation in wide unsupported areas and merging of close features) compromise the quality of pattern transfer. We show that ultrathin (

This work focuses on the development of electrochemical impedance biosensors based on capacitance readout, for the detection of biomolecules in small sample volumes. We performed electrochemical impedance spectroscopy (EIS) measurements of DNA hybridization in electrochemical cells with microfabricated gold electrodes. The time stability of the device was tested in two different configurations: two microelectrodes in a microfluidic channel; two microelectrodes plus a reference electrode in an electrochemical cell. Our results demonstrate that the three-electrode setup is more stable, more reproducible, and suitable for real-time measurements. In the last part of the work we perform a test study of DNA hybridization in real time, and we show that the three-electrode configuration can measure the process in situ and in real time. (C) 2013 Elsevier B.V. All rights reserved.

A technological scheme for the fabrication of metallic point contacts is presented. Nanoimprint lithography (NIL) (J. Vac. Sci. Technol. B, 14(6) (1996) 4129–4133) plays a key role in the process determining the critical dimension of the point contact. The pattern obtained by imprint has been studied by scanning electron microscopy (SEM) and atomic force microscopy (AFM). The masking properties of the polymer film which also serves as an insulating layer in our device, separating the electrodes between which the point contact is located, have been controlled. Our study confirms that the imprint step involved in the process and the chosen thermoplastic polymer meet the requirements for fabricating nm-scale metallic point contacts.

The exposed surface area of porous materials is usually determined by measuring the mass of adsorbed gas as a function of vapour pressure. Here we report a comprehensive study of adsorption in systems with closed bottom, not interconnected pores exhibiting different degrees of disorder, produced with methods encompassing nanolithography and dry and wet etching. Detailed adsorption studies of these matrices show hysteresis loops, as found always in pores having sizes of tens to hundreds of nanometres. The observed variations in the loop shape are associated with changes in the pore morphology. In regular pores formed by vertical and smooth walls, continuous adsorption is found for the first time in agreement with thermodynamic considerations valid for ideal pores. This suggests that irregularities in the walls and pore openings are the key factors behind the hysteresis phenomenon. Interestingly, pores having rough walls but a pyramidal shape also do not show any hysteresis.

In this work the fabrication and characterization of boron-doped diamond (BDD) nanoelectrode arrays are discussed. The use of boron-doped diamond electrodes is very attractive due to advantageous properties including high reproducibility, stability, and robustness under extreme conditions, where conventional electrode materials may undergo severe erosion. BDD electrodes have also proved to be very useful because they show an extremely wide potential window in aqueous solutions without oxidation of the electrode itself. This allows electrochemical detection, at tiny background currents, of a number of substances that oxidize at very positive potentials, where other electrodic materials are not suitable. BDD based NEAs were prepared using Si < 100 > substrates coated with a layer of Boron doped diamond as macroelectrode. NEAs were obtained by creating an array of nanoholes by electron beam lithography (EBL) in a thin film of polycarbonate deposited on top of the macroelectrode. This approach leads to the formation of recessed nanoelectrodes. The parameters for using polycarbonate as a novel electron beam resist have been optimized and successfully used for fabrication of NEAs. The most interesting properties of this polymer for nanofabrication purposes are the high lithographic contrast, which allows the creation of structures of dimensions less than 100 nm; chemical stability, which guarantees a long-term use in electrochemical solutions and the possibility of functionalization with biological molecules (DNA and proteins). NEAs have been characterized with cyclic voltammetry and have provided voltammetric signals controlled by pure radial diffusion. The low background current of BDD added to the properties of NEAs indicate that this system can be applied for the development of sensors with high sensitivity. Polycarbonate surface of NEAs was successfully functionalized with small ss-DNA sequence, confirming the possibility of exploiting these systems as diagnostic biosensors. (C) 2013 Elsevier B.V. All rights reserved.

A new, unique spin-on metal-organic resist presenting a combination of worthwhile properties is described by Giovanna Brusatin and co-workers on page 6261. The resist shows good processability, typical of polymeric resists, direct patternability with various forms of radiation, mild post-exposure treatment requirements, positive or negative tone development, and a selectivity greater than 100:1 with respect to the underlying silicon in a fluorine-based continuous plasma-etching process. The use of such materials for high-resolution deep silicon etching is time- and cost-effective compared with the pattern-transfer materials that are generally used.

We demonstrate that ultrathin (thickness below 1 μm) elastomeric stamps, directly fabricated on rigid substrates can be reliably used in microcontact printing (μCP), showing enhanced mechanical properties with respect to the thicker ones commonly utilized. Macroscopic distorsion (due to thermal expansion and external stress) and microscopic effects such as the membrane relaxation in wide unsupported areas are drastically reduced. A mix and match process (optical lithography followed by μCP) performed with submicron overlay accuracy proves the reliability of the present approach and its applicability to multilevel production processes.

Mechanical oscillators became a main focus of research in recent years for potential applications in biomolecule detectors. We recently demonstrated the feasibility of a scheme based on twin cantilevers with a sensitivity down to the single molecule. This approach is extremely promising under the condition that the two terminals of the device can be functionalized with high selectivity and nanometric accuracy by linker molecules. Here we demonstrate a chemo-mechanical method to achieve the intrinsically aligned functionalization of two silicon surfaces, which can be separated by a gap controllable with nanometric precision. The chemical binding of the target molecules in the selected position is obtained through a cycloaddition reaction which exploits the reactivity of the freshly cleaved surfaces that form when the cantilever gap is created. The general validity of this approach is shown by the use in different chemical environments of two compounds with different reactive functional groups.

Embryonic stem (ES) cell differentiation in specific cell lineages is a major issue in cell biology particu- larly in regenerative medicine. Differentiation is usually achieved by using biochemical factors and it is not clear whether mechanical properties of the substrate over which cells are grown can affect proliferation and differentiation. Therefore, we produced patterns in polydimethylsiloxane (PDMS) consisting of groove and pillar arrays of sub- micrometric lateral resolution as substrates for cell cultures. We analyzed the effect of different nanostructures on dif- ferentiation of ES-derived neuronal precursors into neuro- nal lineage without adding biochemical factors. Neuronal precursors adhered on PDMS more effectively than on glass coverslips. We demonstrated that neuronal yield was en- hanced by increasing pillars height from 35 to 400 nm. On higher pillar neuronal differentiation reaches � 80% 96 h after plating and the largest differentiation enhancement of pillars over flat PDMS was observed during the first 6 h of culture. We conclude that PDMS nanopillars accelerate and increase neuronal differentiation. Biotechnol. Bioeng. 2011;108: 2736-2746. 2011 Wiley Periodicals, Inc.

Ordered arrays of nanoelectrodes for electrochemical use are prepared by electron beam lithography (EBL) using polycarbonate as a novel e-beam resist. The nanoelectrodes are fabricated by patterning arrays of holes in a thin film of polycarbonate spin-coated on a gold layer on Si/Si(3)N(4) substrate. Experimental parameters for the successful use of polycarbonate as high resolution EBL resist are optimized. The holes can be filled partially or completely by electrochemical deposition of gold. This enables the preparation of arrays of nanoelectrodes with different recession degree and geometrical characteristics. The polycarbonate is kept on-site and used as the insulator that separates the nanoelectrodes. The obtained nanoelectrode arrays (NEAs) exhibit steady state current controlled by pure radial diffusion in cyclic voltammetry for scan rates up to approximately 50 mV s( - 1). Electrochemical results showed satisfactory agreement between experimental voltammograms and suitable theoretical models. Finally, the peculiarities of NEAs versus ensembles of nanoelectrodes, obtained by membrane template synthesis, are critically evaluated.

A positive tone resist for UV and X-ray lithography synthesized starting from an organically modified silicon alkoxide, bis(triethoxysilyl)benzene, through the sol–gel method, either in basic or in acid catalysis, is presented. Being directly photo-processable, the sol–gel system combines the opportunity to avoid the use of a sacrificial layer in the fabrication process, with the possibility to fit electro-optical and structural properties of the final device material to specific requirements. In addition, the positive tone behaviour allows to preserve the organic functionality of the system after irradiation. A study of the optical and structural modifications induced on the resist by irradiation has been carried out by FT-IR spectroscopy, UV–vis spectroscopy and spectroscopic ellipsometry. An interpretation of the mechanisms leading to exposed cross-linked film development is given. Experiments have demonstrated the possibility of obtaining structures on films with lateral dimensions spanning from the micron scale up to less than a hundred nm, opening the way to a possible exploitation of such positive tone functional system in the field of miniaturized sensors.

Nanoelectrochemical immunosensors fabricated by templated electrodeposition of gold nanoelectrodes inside the pores of polycarbonate (PC) track-etched membranes, followed by the immobilization of the biorecognition elements on the surrounding PC, have proven high sensitivity and specificity for protein detection. The signal transduction scheme involves a suitable redox mediator added to the sample solution to shuttle electrons from the gold nanoelectrodes to the biorecognition layer, both elements being in strict spatial proximity. Highly improved signal-to-background current ratio, which are peculiar of NEEs with respect to other electrochemical transducers, can be exploited in this way. Two detection schemes were tested: one based on the direct immobilization of the target protein on the PC of the NEE (approach A) and the other based on the immobilisation on PC of an antibody to capture the target protein (approach B). The biorecognition process was completed by adding a primary antibody and a secondary antibody with horse radish peroxidase (HRP) as enzyme label; methylene blue was the redox mediator added to the electrolyte solution. Typical target analytes were single chain fragment variable proteins, for approach A, and trastuzumab (also known as Herceptin®), for approach B. NEE-based capture sensors were tested successfully to detect small amounts of the receptor protein HER2 in biological samples. Finally, motivated by the target of a better control of the geometrical characteristics of ensembles of nanoelectrodes (size, density, geometrical arrangement, and degree of recession), and by the positive results obtained with track-etch membranes of PC from the standpoint of protein immobilization, we demonstrated the fabrication of nanobiosensors by patterning ordered arrays of nanoelectrodes (NEAs) by electron beam lithography (EBL) on polycarbonate. EBL results perfectly suitable for the top-down fabrication of arrays of nanobiosensors on thin PC films deposited on gold coated silicon.

We propose fast nanoimprinting lithography (NIL) process based on the use of stamps with integrated heater. The latter consists of heavily ion implantation n-type doped silicon layer buried below the microstructured surface of the stamp. The stamp is heated by Joule effect, by 50 mu s 25 Hz repetition rate current pulses flowing in the conductive layer. Using this approach we have reproducibly imprinted areas of similar to 2 cm(2) within 16 s with residual layers in the range of few tens of nm. This result paves the way for processes in the sub-1 s timescale over large area surfaces. (C) 2008 Elsevier B.V. All rights reserved.

"in this study we demonstrate the possibility to prepare highly sensitive nanostructured electrochemical immunosensors by immobilizing biorecognition elements on nanoelectrode ensembles (NEEs) prepared in track-etch polycarbonate membranes. The gold nanodisk electrodes act as electrochemical transducers while the surrounding polycarbonate binds the antibody-based biorecognition layer. The interaction between target protein and antibody is detected by suitable secondary antibodies labelled with a redox enzyme. A redox mediator, added to the sample solution, shuttles electrons from the nanoelectrodes to the biorecognition layer, so generating an electrocatalytic signal. This allows one to fully exploit the highly improved signal-to-background current ratio, typical of NEEs. In particular, the receptor protein HER2 was studied as the target analyte. HER2 detection allows the identification of breast cancer that can be treated with the monoclonal antibody trastuzumab. NEEs were functionalized with trastuzumab which interacts specifically with HER2. The biorecognition process was completed by adding a primary antibody and a secondary antibody labelled with horseradish peroxidase. Hydrogen peroxide was added to modulate the label electroactivity; methylene blue was the redox mediator generating voltammetric signals. NEEs functionalized with trastuzumab were tested to detect small amounts of HER2 in diluted cell lysates and tumour lysates. (c) 2008 Elsevier B.V. All rights reserved."

A new method for preparing 3D scaffolds for tissue engineering applications with highly controlled micro and nanotopography have been developed. A combination of UV and electron beam lithography was employed for master fabrication and nanoimprint lithography for the preparation of the 3D polymeric scaffolds.

This review discusses some of the most common polymer scaffold fabrication techniques used for tissue engineering applications. Although the field of scaffold fabrication is now well established and advancing at a fast rate, more progress remains to be made, especially in engineering small diameter blood vessels and providing scaffolds that can support deep tissue structures. With this in mind, we introduce two new lithographic methods that we expect to go some way to addressing this problem.

The objective of this benchmarking is to establish a comparison of several tools and processes used in thermal NIL with Si stamps at the nanoscale among the authors' laboratories. The Si stamps have large arrays of 50 nm dense lines and were imprinted in all these laboratories in a similar to 100 nm thick mr-18010E film. Other materials, such as mr-17010E, were also tested. Good patterns were obtained and some limitations were identified. Reducing the pressure to 15 bars enables the printing of 50 nm structures without pulling them off. At higher pressures, some bending effects resulting in pattern deformation were observed. It was proven that a pressure of 1.5 bars is sufficient to imprint perfect 50 nm lines. The influence of the antiadhesive layer and mold design has been characterized by the demonstration of pulled off lines in some cases. Moreover, it has been shown that the scatterometry method is particularly useful for the characterization of 50 nm lines and that the residual layer thickness corresponds to the theoretical estimate as long as the lines are well defined. One process was demonstrated which combines high reproducibility with high throughput, achieving a cycle time of 2 min. (c) 2007 American Vacuum Society.

One of the topics of the biomedical research is to design and develop new drug delivery systems able to perform time-controlled release of medicals with target specificity. Porous silicon, characterized by macropores of 2 mu m average diameter, with a 200 nm thick nanoporous coverage, was produced and its potential applications as drug-carrier were investigated. Doxorubicin loaded porous silicon bits exhibited a time-depending drug delivery profile that is in trend with the cytotoxicity of adenocarcinoma cancer cells exposed to bits. (c) 2006 Published by Elsevier B.V.

We introduce simple double-casting replication methods for high-aspect-ratio microstructures fabricated by deep x-ray lithography using intermediate molds of soft materials. Two types of soft material are investigated. The ability to fabricate polymethylsiloxane (PDMS) molds with well-type structures with aspect ratios up to 35: 1 is demonstrated for structure densities below 50%, and the reproduction from this mold of pillar-type polymethyl methacrylate (PMMA) structures with aspect ratios of 20: 1 is achieved. Polyvinyl alcohol (PVA), a water soluble polymer, is also tested as a sacrificial intermediate mold and successfully used for the replication of structures with aspect ratios up to 5:1. Double-casting replication methods are described and discussed for their potential improvement. (c) 2006 Society of Photo-Optical Instrumentation Engineers.

Nanotechnology is rapidly opening new possibilities in a wide variety of disciplines. Biomedical micro- and nano-devices, with components geometries as large as hundreds of microns to as small as a few nanometers, have potential uses that range from the analysis of biomolecules to disease diagnosis, prevention and treatment. Exciting technological advances have been made in nanofabrication and in micro- and nano-electromechanical systems [1] (MEMS, NEMS), microfluidic devices [2], micro- or nano-optics (diffractive optics [3], high efficiency multilevel zone plates [4], photonic crystals [5]), and innovations in the toolset of biology (microfluidic chips for DNA array [6]) and medical (microsurgical tools [7] and drug delivery), provide examples of advancement in nanotechnology. The application of carbon nanotube atomic force microscopy (AFM) imaging enabled multiplex detection of the positions of specific DNA sequences labelled with nanometer-size tags [8]. This method allowed the direct determination of haplotypes of patient samples, and provided a more accurate assessment of cancer risk from the UGT1A7 gene [8]. The phage display approach for isolating peptides with high binding affinities to specific crystal types discussed by Whaley et al. [9] might find use both in synthesis and self-assembly of hetero-structures made from nano-particles. The research in tissue engineering stands to benefit most from our growing ability to fabricate complex nano-structured materials. The enormous potential in nanotechnology is at least a few years from beginning to materialize the products and technologies involved in the medical application of nanotechnology. But the possibility of improved speed, greater sensitivity, reduced cost, and decreased invasiveness, has generated substantial interest in miniaturized devices.

We present results on a lithographic approach that combines nanoimprint (NIL) and x-ray lithography (XRL) for fabricating unconventional three-dimensional (3D) polymer structures. The use of XRL for structuring a prepatterned resist by NIL gives rise to high-resolution high-aspect-ratio structures whose overall profile is enveloped by the original 3D imprinted profile. The technological potential of this method has been demonstrated by patterning several different types of structures with XRL on an hexagonal array of hemispheres previously obtained by nanoimprinting.

Microcontact printing (μCP) and Nanoimprint lithography (NIL) have both proven to be high resolution lithography processes suitable for large area parallel lithography.1–7 This property makes them both promising for application in device fabrication, where high throughput is an issue. Especially NIL is used by numerous groups due to its high resolution and potential for applications.8–10

We describe a new approach of microcontact printing which is based on a tri-layer processing. The replication process has four steps: (1) preparation of gold-PMMA-substrate by sputtering and spincoating deposition; (2) pattern transfer with alkanethiol molecules from a PDMS stamp to gold; (3) wet etching of gold and (4) reactive ion etching of under-layer PMMA. By this method, we succeeded in fabricating microstructures in PMMA resist which are suitable for further hard material pattern transfers, thereby providing an enhanced process compatibility with most of microfabrication steps

Embryonic stem (ES) cell differentiation in specific cell lineage is a major issue in cell biology particularly in regenerative medicine. Differentiation is usually achieved by using biochemical factors which concentration and mechanism are not completely understood and with sides effects difficult to overcome. Using a substrate which mimics brain extracellular matrix it could be possible to induce ES-cells differentiation into neurons without adding any biochemical factors. Therefore, we produced patterns in polydimethylsiloxane (PDMS) consisting of groove and pillar arrays of sub- micrometric lateral resolution as substrates for cell cultures. Neuronal precursors from ES cells were obtained using a Stromal Cell- Derived Inducing Activity protocol and we analyzed the effect of different nanostructures on differentiation into neuronal lineage. Neuronal precursors adhered on PDMS more effectively than on glass coverslips. After 48 hours of culture on PDMS pillars with a 500nm period, neuronal differentiation increased and almost doubled with respect to flat PDMS substrates. Neuronal yield was enhanced by increasing pillars height from 35 to 400 nm. With pillars, 500nm period and 360nm height, the neuronal yield reached ∼80% 96 hours after plating. However the largest differentiation enhancement of pillars over flat PDMS was observed during the first 6 hours of culture. These shown results that PDMS nanopillars accelerate ES cells differentiation into neurons.

Recently, there has been growing interest in sensors based on frequency shift detection of mechanical resonators, that has led to extremely sensitive diagnostic methods for medicine and biology. Here the authors propose and demonstrate a strategy capable of detecting the presence of a few macromolecules, which uses an asymmetrical cantilever structure of several microns in size, operated at room temperature, and under ordinary vacuum conditions. The idea is to detect the presence of one or more molecules by detecting the mechanical cross-talk induced by the molecular link between a short cantilever and an array of longer cantilevers facing the short one. (c) 2007 American Institute of Physics.

The repair of vascular tubing is an important task in tissue engineering. The behavior of cells is strongly influenced by the topology of the surfaces, on both a micrometric and a nanometric scale, in their vicinity. Thus the authors wish to make tubes that are patterned on the inner surface. One way to do this is to use the good depth of focus capabilities of x-ray exposure to print an array of dots, 200 nm diameter and 400 nm pitch, onto a curved surface coated in resist. A die made from this structure allows nanoembossing into a biodegradable polymer. A closed vessel can then be made by adding a lid, that also has a similar nanopatterned surface. Details of the accuracy of transfer are given, It is concluded that x-ray printing is a suitable approach for the formation of internally patterned tubing. (c) 2006 American Vacuum Society.

In nanoimprint lithography, high pressures are utilised for embossing thermoplastics. This may introduce mechanical strain in the mould and the sample. By using a thermocurable polymer (polydimethylsiloxane, PDMS), a low pressure imprint process, working at pressures of < 0.2 MPa, has proven viable. The new process may drastically reduce the thermal budget and the mechanical load in imprint lithography.

A new patterning process based on the microcontact printing of self-assembled monolayers on gold is presented. The method is readily adapted to processes utilising standard optical or electron beam resists, and is fully compatible with industrial semiconductor fabrication technology. The process can also be applied to surfaces which exhibit extreme nonplanarity such as sharp edges of mesas or etch grooves.