Steven Lenhert

Publications of Steven Lenhert

• Multifunctional lipid multilayer stamping.

  Authors: Omkar A Nafday, Troy W Lowry, Steven Lenhert

  Small (Weinheim an der Bergstrasse, Germany). 02/2012; 8(7):1021-8.

  Nanostructured lipid multilayers on surfaces are a promising biofunctional nanomaterial. For example, surface-supported lipid multilayer diffraction gratings with optical properties that depend onNanostructured lipid multilayers on surfaces are a promising biofunctional nanomaterial. For example, surface-supported lipid multilayer diffraction gratings with optical properties that depend on the microscale spacing of the grating lines and the nanometer thickness of the lipid multilayers have been fabricated previously by dip-pen nanolithography (DPN), with immediate applications as label-free biosensors. The innate biocompatibility of such gratings makes them promising as biological sensor elements, model cellular systems, and construction materials for nanotechnology. Here a method is described that combines the lateral patterning capabilities and scalability of microcontact printing with the topographical control of nanoimprint lithography and the multimaterial integration aspects of dip-pen nanolithography in order to create nanostructured lipid multilayer arrays. This approach is denoted multilayer stamping. The distinguishing characteristic of this method is that it allows control of the lipid multilayer thickness, which is a crucial nanoscale dimension that determines the optical properties of lipid multilayer nanostructures. The ability to integrate multiple lipid materials on the same surface is also demonstrated by multi-ink spotting onto a polydimethoxysilane stamp, as well as higher-throughput patterning (on the order of 2 cm(2) s(-1) for grating fabrication) and the ability to pattern lipid materials that could not previously be patterned with high resolution by lipid DPN, for example, the gel-phase phospholipid 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) or the steroid cholesterol.

• High-throughput optical quality control of lipid multilayers fabricated by dip-pen nanolithography.

  Authors: Omkar A Nafday, Steven Lenhert

  Nanotechnology. 06/2011; 22(22):225301.

  Surface supported phospholipid multilayers are promising materials for nanotechnology because of their tendency to self-organize, their innate biocompatibility, the possibility to encapsulate otherSurface supported phospholipid multilayers are promising materials for nanotechnology because of their tendency to self-organize, their innate biocompatibility, the possibility to encapsulate other materials within the multilayers, and the ability to control the multilayer thickness between ∼ 2 and 100 nm during fabrication. Dip-pen nanolithography (DPN) is an atomic force microscopy (AFM) based fabrication method that allows high-throughput fabrication and integration of a variety of micro- and nanostructured materials including lipid multilayers, with areal throughputs on the scale of cm(2) min(-1). Although multilayer thickness is a critical feature that determines the functionality of the lipid multilayer structures (for instance as carriers for other materials as well as optical scattering properties), reliable height characterization by AFM is slow (on the order of µm(2) min(-1)) and a bottleneck in the lithographic process. Here we describe a novel optical method to reliably measure the height of fluorescent multilayers with thicknesses above 10 nm, and widths above the optical diffraction limit based on calibrating the fluorescence intensity using one-time AFM height measurements. This allows large surface areas to be rapidly and quantitatively characterized using a standard fluorescence microscope. Importantly, different pattern dimensions (0D dots, 1D lines or 2D squares) require different calibration parameters, indicating that shape influences the optical properties of the structured lipid multilayers. This method has general implications in the systematic and high-throughput optical characterization of nanostructure-function relationships.

• In situ lipid dip-pen nanolithography under water.

  Authors: Steven Lenhert, Chad A Mirkin, Harald Fuchs

  Scanning. 03/2010; 32(1):15-23.

  Lipids form the structural and functional basis of biological membranes, and methods for studying their self-organization in well-defined nano- and micro-scale model systems can provide insights intoLipids form the structural and functional basis of biological membranes, and methods for studying their self-organization in well-defined nano- and micro-scale model systems can provide insights into biology. Using lipids as an ink for dip-pen nanolithography (lipid DPN) permits the rapid nanostructuring of multicomponent model lipid membrane systems, but this technique has so far been limited to air. Here we demonstrate that lipid DPN can be carried out under water with single tips or parallel arrays. Using the same tip for deposition and imaging in aqueous solution permits imaging of self-spreading lipid bilayer spots in situ and quantification of the nanoscale spreading kinetics in real time by means of lateral-force microscopy. Furthermore, using fluorophore-labeled phospholipids, we directly observed, by confocal laser scanning microscopy, a two-phase (oil in water) meniscus formed around the contact point between the DPN tip and surface, gaining insights into the mechanisms of the ink transport. The methods described here provide a new tool and environment for high-resolution studies of lipid nanodynamics and molecular printing processes in general.

• Common genetic pathways regulate organ-specific infection-related development in the rice blast fungus.

  Authors: Sara L Tucker, Maria I Besi, Rita Galhano, Marina Franceschetti, Stephan Goetz, Steven Lenhert, Anne Osbourn, Ane Sesma

  The Plant cell. 03/2010; 22(3):953-72.

  Magnaporthe oryzae is the most important fungal pathogen of rice (Oryza sativa). Under laboratory conditions, it is able to colonize both aerial and underground plant organs using differentMagnaporthe oryzae is the most important fungal pathogen of rice (Oryza sativa). Under laboratory conditions, it is able to colonize both aerial and underground plant organs using different mechanisms. Here, we characterize an infection-related development in M. oryzae produced on hydrophilic polystyrene (PHIL-PS) and on roots. We show that fungal spores develop preinvasive hyphae (pre-IH) from hyphopodia (root penetration structures) or germ tubes and that pre-IH also enter root cells. Changes in fungal cell wall structure accompanying pre-IH are seen on both artificial and root surfaces. Using characterized mutants, we show that the PMK1 (for pathogenicity mitogen-activated protein kinase 1) pathway is required for pre-IH development. Twenty mutants with altered pre-IH differentiation on PHIL-PS identified from an insertional library of 2885 M. oryzae T-DNA transformants were found to be defective in pathogenicity. The phenotypic analysis of these mutants revealed that appressorium, hyphopodium, and pre-IH formation are genetically linked fungal developmental processes. We further characterized one of these mutants, M1373, which lacked the M. oryzae ortholog of exportin-5/Msn5p (EXP5). Mutants lacking EXP5 were much less virulent on roots, suggesting an important involvement of proteins and/or RNAs transported by EXP5 during M. oryzae root infection.

• Lipid multilayer gratings.

  Authors: Steven Lenhert, Falko Brinkmann, Thomas Laue, Stefan Walheim, Christoph Vannahme, Soenke Klinkhammer, Miao Xu, Sylwia Sekula, Timo Mappes, Thomas Schimmel, Harald Fuchs

  Nature nanotechnology. 02/2010; 5(4):275-9.

  The interaction of electromagnetic waves with matter can be controlled by structuring the matter on the scale of the wavelength of light, and various photonic components have been made by structuringThe interaction of electromagnetic waves with matter can be controlled by structuring the matter on the scale of the wavelength of light, and various photonic components have been made by structuring materials using top-down or bottom-up approaches. Dip-pen nanolithography is a scanning-probe-based fabrication technique that can be used to deposit materials on surfaces with high resolution and, when carried out in parallel, with high throughput. Here, we show that lyotropic optical diffraction gratings--composed of biofunctional lipid multilayers with controllable heights between approximately 5 and 100 nm--can be fabricated by lipid dip-pen nanolithography. Multiple materials can be simultaneously written into arbitrary patterns on pre-structured surfaces to generate complex structures and devices, allowing nanostructures to be interfaced by combinations of top-down and bottom-up fabrication methods. We also show that fluid and biocompatible lipid multilayer gratings allow label-free and specific detection of lipid-protein interactions in solution. This biosensing capability takes advantage of the adhesion properties of the phospholipid superstructures and the changes in the size and shape of the grating elements that take place in response to analyte binding.

• Measurement of DPN-Ink Viscosity using an AFM Cantilever

  Authors: Soma Biswas, Michael Hirtz, Steven Lenhert, Harald Fuchs

  Nanotechnology 2010: Electronics, Devices, Fabrication, MEMS, Fluidics and Computational; 01/2010

  The Dip-Pen Nanolithography (DPN) process uses a chemically coated scanning probe tip (the “pen”) to directly deposit a material (“ink”) with nanometer precision onto a substrate. SeveralThe Dip-Pen Nanolithography (DPN) process uses a chemically coated scanning probe tip (the “pen”) to directly deposit a material (“ink”) with nanometer precision onto a substrate. Several experimental parameters have been observed to influence the ink transport in DPN. Among these, viscosity and density of the ink are the two important parameters which play a crucial role in ink transport in DPN. It is necessary to understand and control these parameters in order to optimize DPN processes for a particular application. Here we report on studies aimed at employing the atomic force microscope (AFM) to measure the viscosity of ink used for DPN. Generally, for DPN, the ink wells are filled with ink (biological molecules in chloroform) and the ink then gets transported to the micro channels from the ink reservoir. The ink wells are dried for few hours to allow for evaporating of the chloroform before using them for coating the tip. It is difficult to measure the viscosity of this ink using standard p

• Multiplexed Lipid Dip-Pen Nanolithography on Subcellular Scales for the Templating of Functional Proteins and Cell Culture.

  Authors: Sylwia Sekula, Jeanette Fuchs, Susanne Weg-Remers, Peter Nagel, Stefan Schuppler, Joe Fragala, Nora Theilacker, Matthias Franzreb, Christer Wingren, Peter Ellmark, Carl A K Borrebaeck, Chad A Mirkin, Harald Fuchs, Steven Lenhert

  Small (Weinheim an der Bergstrasse, Germany). 10/2008;

  Molecular patterning processes taking place in biological systems are challenging to study in vivo because of their dynamic behavior, subcellular size, and high degree of complexity. In vitroMolecular patterning processes taking place in biological systems are challenging to study in vivo because of their dynamic behavior, subcellular size, and high degree of complexity. In vitro patterning of biomolecules using nanolithography allows simplification of the processes and detailed study of the dynamic interactions. Parallel dip-pen nanolithography (DPN) is uniquely capable of integrating functional biomolecules on subcellular length scales due to its constructive nature, high resolution, and high throughput. Phospholipids are particularly well suited as inks for DPN since a variety of different functional lipids can be readily patterned in parallel. Here DPN is used to spatially pattern multicomponent micro- and nanostructured supported lipid membranes and multilayers that are fluid and contain various amounts of biotin and/or nitrilotriacetic acid functional groups. The patterns are characterized by fluorescence microscopy and photoemission electron microscopy. Selective adsorption of functionalized or recombinant proteins based on streptavidin or histidine-tag coupling enables the semisynthetic fabrication of model peripheral membrane bound proteins. The biomimetic membrane patterns formed in this way are then used as substrates for cell culture, as demonstrated by the selective adhesion and activation of T-cells.

• A self-correcting inking strategy for cantilever arrays addressed by an inkjet printer and used for dip-pen nanolithography.

  Authors: Yuhuang Wang, Louise R Giam, Matt Park, Steven Lenhert, Harald Fuchs, Chad A Mirkin

  Small (Weinheim an der Bergstrasse, Germany). 10/2008; 4(10):1666-70.

• Probing the organization of photosystem II in photosynthetic membranes by atomic force microscopy.

  Authors: Helmut Kirchhoff, Steven Lenhert, Claudia Büchel, Lifeng Chi, Jon Nield

  Biochemistry. 02/2008; 47(1):431-40.

  Efficient photosynthetic energy transduction and its regulation depend on a precise supramolecular arrangement of the plant photosystem II (PSII) complex in grana membranes of chloroplasts. TheEfficient photosynthetic energy transduction and its regulation depend on a precise supramolecular arrangement of the plant photosystem II (PSII) complex in grana membranes of chloroplasts. The topography of isolated photosystem II supercomplexes and the supramolecular organization of this complex in grana membrane preparations are visualized by high-resolution atomic force microscopy (AFM) in air in tapping mode with an active feedback control to minimize tip-sample interactions. Systematic comparison between topographic characteristics of the protrusions in atomic force microscopic images and well-established high-resolution and freeze-fracture electron microscopic data shows that the photosystem II organization can be properly imaged by AFM in air. Taking the protruding water-splitting apparatus as a topographic marker for PSII, its distribution and orientation in isolated grana membrane were analyzed. For the latter a new mathematical procedure was established, which revealed a preference for a parallel alignment of PSII that resembles the organization in highly ordered semicrystalline arrays. Furthermore, by analyzing the height of grana membrane stacks, we conclude that lumenal protrusions of adjacent photosystem II complexes in opposing membranes are displaced relative to each other. The functional consequences for lateral migration processes are discussed.

• Capillary-induced contact guidance.

  Authors: Steven Lenhert, Ane Sesma, Michael Hirtz, Lifeng Chi, Harald Fuchs, Hans-Peter Wiesmann, Anne E Osbourn, Bruno M Moerschbacher

  Langmuir : the ACS journal of surfaces and colloids. 10/2007; 23(20):10216-23.

  Topographical features are known to impose capillary forces on liquid droplets, and this phenomenon is exploited in applications such as printing, coatings, textiles and microfluidics. SurfaceTopographical features are known to impose capillary forces on liquid droplets, and this phenomenon is exploited in applications such as printing, coatings, textiles and microfluidics. Surface topographies also influence the behavior of biological cells (i.e., contact guidance), with implications ranging from medicine to agriculture. An accurate physical description of how cells detect and respond to surface topographies is necessary in order to move beyond a purely heuristic approach to optimizing the topographies of biomaterial interfaces. Here, we have used a combination of Langmuir-Blodgett lithography and nanoimprinting to generate a range of synthetic microstructured surfaces with grooves of subcellular dimensions in order to investigate the influence of capillary forces on the biological process of contact guidance. The physical-chemical properties of these surfaces were assessed by measuring the anisotropic spreading of sessile water droplets. Having established the physical properties of each surface, we then investigated the influence of capillary forces on the processes of cellular contact guidance in biological organisms, using mammalian osteoblasts and germinating fungal spores as tester organisms. Our results demonstrate that capillary effects are present in topographical contact guidance and should therefore be considered in any physical model that seeks to predict how cells will respond to a particular surface topography.

• Langmuir-Blodgett patterning: a bottom-up way to build mesostructures over large areas.

  Authors: Xiaodong Chen, Steven Lenhert, Michael Hirtz, Nan Lu, Harald Fuchs, Lifeng Chi

  Accounts of chemical research. 07/2007; 40(6):393-401.

  This Account describes a new paradigm, Langmuir-Blodgett (LB) patterning, for large-area patterning with mesostructured features based on the well-established LB technique. This strategy uses aThis Account describes a new paradigm, Langmuir-Blodgett (LB) patterning, for large-area patterning with mesostructured features based on the well-established LB technique. This strategy uses a simple fabrication technique to control the alignment, size, shape, and periodicity of self-organized phospholipid monolayer patterns with feature sizes down to 100 nm over surface areas of square centimeters. Because of the anisotropic wetting behavior of the patterns, they can be used as templates to direct the self-assembly of functional molecules and nanocrystals. Furthermore, the chemical patterns can be converted into topographical structures, which can be used to direct cell growth and organize nanocrystals. The mesoscopic structured surfaces described here may serve as a platform in engineering the biological/material interface and constructing biofunctionalized structures and "programmed" systems.

• Massively parallel dip-pen nanolithography of heterogeneous supported phospholipid multilayer patterns.

  Authors: Steven Lenhert, Peng Sun, Yuhuang Wang, Harald Fuchs, Chad A Mirkin

  Small (Weinheim an der Bergstrasse, Germany). 02/2007; 3(1):71-5.

• Interface interaction controlled transport of CdTe nanoparticles in the microcontact printing process.

  Authors: Xiaochun Wu, Steven Lenhert, Lifeng Chi, Harald Fuchs

  Langmuir : the ACS journal of surfaces and colloids. 09/2006; 22(18):7807-11.

  High-quality CdTe nanoparticles stabilized with thioglycolic acid (TGA) are patterned on SiO2/Si surfaces using microcontact printing (microCP). Due to the weak interaction of the nanoparticles withHigh-quality CdTe nanoparticles stabilized with thioglycolic acid (TGA) are patterned on SiO2/Si surfaces using microcontact printing (microCP). Due to the weak interaction of the nanoparticles with the stamp surface, tailoring of gas flow rate during the inking process as well as the type and scale of the patterns on the stamp are used to control the distribution of the nanoparticles on the structured stamp surface. This distribution is then transferred the printed regions. Either edge printing or homogeneous printing can be achieved under optimized conditions. In addition, new structures such as nanowires form under certain conditions.

• Mechanism of regular pattern formation in reactive dewetting.

  Authors: Steven Lenhert, Michael Gleiche, Harald Fuchs, Lifeng Chi

  Chemphyschem : a European journal of chemical physics and physical chemistry. 01/2006; 6(12):2495-8.

• Osteoblast alignment, elongation and migration on grooved polystyrene surfaces patterned by Langmuir-Blodgett lithography.

  Authors: Steven Lenhert, Marie-Beatrice Meier, Ulrich Meyer, Lifeng Chi, Hans-Peter Wiesmann

  Biomaterials. 03/2005; 26(5):563-70.

  Topographically patterned surfaces are known to influence cellular behavior in a controllable manner. However, the relatively large surface areas (several cm2) required for many biomaterialTopographically patterned surfaces are known to influence cellular behavior in a controllable manner. However, the relatively large surface areas (several cm2) required for many biomaterial applications are beyond the practical limits of traditional lithography. Langmuir-Blodgett lithography, a recently developed method, was used to fabricate regularly spaced grooves of different depths (50 and 150 nm) with a periodicity of 500 nm over several square centimeter on silicon surfaces. These topographies were transferred into polystyrene surfaces by means of nanoimprinting. Primary osteoblasts were cultured on the patterned polymer surfaces. They were observed to align, elongate and migrate parallel to the grooves. The combination of Langmuir-Blodgett lithography with nanoimprinting enables the fabrication of large, nanostructured surface areas on a wide spectrum of different biomaterials. Osteoblasts show a significant anisotropic behavior to these surfaces, which can enhance cell settlement on the surface or be used to direct tissue generation on the biomaterial interface.

• Transversal and lateral exciton energy transfer in grana thylakoids of spinach.

  Authors: Helmut Kirchhoff, Mauricio Borinski, Steven Lenhert, Lifeng Chi, Claudia Büchel

  Biochemistry. 12/2004; 43(45):14508-16.

  The excitation energy transfer between photosystem (PS) II complexes was studied in isolated grana disks and thylakoids using chlorophyll a fluorescence induction measurements in the presence of DCMUThe excitation energy transfer between photosystem (PS) II complexes was studied in isolated grana disks and thylakoids using chlorophyll a fluorescence induction measurements in the presence of DCMU under stacked and destacked conditions. Destacking of grana was achieved using a sonication protocol in a buffer without MgCl(2). The degree of stacking was controlled and quantified by atomic force microscopy and by the concomitant absorption changes. As expected from the literature, intact thylakoids showed a strong dependency of the connectivity of PSII centers, the F(m)/F(o) ratio as well as the fraction of PSIIbeta centers on the MgCl(2) concentration. In contrast, these parameters did not change in isolated grana disks. In particular, the connectivity remained constantly high irrespective of the degree of destacking. These differences were explained by the high protein density in grana disks, which hinders separation and mixing of proteins sufficiently to change energy transfer properties. Due to the occurrence of stroma lamella in intact thylakoids, intermixing of PSII and PSI is possible and allows for changes in F(m)/F(o) ratio as is the separation of LHCII from PSII, thus leading to an increase in the fraction of PSIIbeta. Even if mixing and separation of proteins are impaired in isolated grana disks, destacking should lead to a decrease in connectivity if transversal excitation energy transfer between two opposite membranes is significant. Because the connectivity is constant over all degrees of destacking employed, we conclude that the energy transfer in granas is mainly lateral.

• Fluorescence excitation on monolithically integrated all-polymer chips.

  Authors: Mauno Schelb, Christoph Vannahme, Alexander Welle, Steven Lenhert, Benjamin Ross, Timo Mappes

  Journal of biomedical optics. 15(4):041517.

  All-polymer chips with integrated optical waveguides and microfluidic channels were built as a platform for fluorescence excitation of biological samples. Their functionality has been shown for fourAll-polymer chips with integrated optical waveguides and microfluidic channels were built as a platform for fluorescence excitation of biological samples. Their functionality has been shown for four cases: (i) fluorescence of labeled phospholipids inside a microfluidic channel, (ii) fluorescence of stained cells inside a microfluidic channel, (iii) fluorescence of labeled phospholipids on top of a polymer waveguide, and (iv) fluorescence of stained cells on top of a polymer waveguide.