Femto‐Second Laser‐Based Free Writing of 3D Protein Microstructures and Micropatterns with Sub‐Micrometer Features: A Study on Voxels,Porosity, and Cytocompatibility

Femto‐second laser‐based free‐writing of complex protein microstructures and micropatterns, with sub‐micrometer features and controllability over voxel dimension, morphology, and porosity, is reported. Protein voxels including lines, spots, and micropillars are fabricated. Laser power, exposure time, z‐position, protein and photosensitizer concentrations, but not scanning speed, are important controlling parameters. A lateral fabrication resolution of ≈200 nm is demonstrated in 2D line voxels. 3D spot voxels are ellipsoids with 400 nm lateral and 1.5 μm axial dimensions. An ascending z‐stack scanning method to verify the theoretical axial optical resolution, delineate and enhance the axial fabrication resolution of 3D structures, including square prism and cylinder micropillars, is also reported. The micropillar array presents a simple “write‐and‐seed” and table platform for cell niche studies. Fibroblasts attach to, grow on, and express adhesion to molecules on micropillar arrays without the need of matrix coating. They exhibit a more “3D” morphology comparing with that in 2D monolayer cultures and physiological functions such as matrix deposition. This work presents an important milestone in engineering complex protein microstructures and micropatterns with sub‐micrometer topological features to mimic the native matrix niche for cell‐matrix interaction studies.     

Surfactant‐Assisted Emulsion Self‐Assembly of Nanoparticles into Hollow Vesicle‐Like Structures and 2D Plates

The emulsion‐based self‐assembly of nanoparticles into low‐dimensional superparticles of hollow vesicle‐like assemblies is reported. Evaporation of the oil phase at relatively low temperatures from nanoparticle‐containing oil‐in‐water emulsion droplets leads to the formation of stable and uniform sub‐micrometer vesicle‐like assembly structures in water. This result is in contrast with those from many previously reported emulsion‐based self‐assembly methods, which produce solid spherical assemblies. It is found that extra surfactants in both the oil and water phases play a key role in stabilizing nanoscale emulsion droplets and capturing hollow assembly structures. Systematic investigation into what controls the morphology in emulsion self‐assembly is carried out, and the approach is extended to fabricate more complex rattle‐like structures and 2D plates. These results demonstrate that the emulsion‐based assembly is not limited to typical thermodynamic spherical assembly structures and can be used to fabricate various types of interesting low‐dimensional assembly structures.     

Gyroid‐Structured 3D ZnO Networks Made by Atomic Layer Deposition

3D continuous ZnO morphologies with characteristic feature sizes on the 10 nm length scale are attractive for electronic device manufacture. However, their synthesis remains a challenge because of the low crystallization temperature of ZnO. Here, we report a method for the robust and reliable synthesis of fully crystalline 3D mesoporous ZnO networks by means of atomic layer deposition (ALD) of ZnO into a self‐assembled block copolymer template. By carefully optimizing the processing conditions we are able to synthesize several‐micrometer‐thick layers of mesoporous ZnO networks with a strut width of 30 nm. Two 3D mesoporous morphologies are manufactured: a periodic gyroid structure and a random worm‐like morphology. Exploiting the ALD property to conformally coat complex surfaces of high aspect ratio, the channel network of a 3D continuous channel network of a self‐assembled block copolymer is replicated into ZnO. X‐ray photoemission spectroscopy and x‐ray diffraction measurements reveal that the chemical composition of the mesoporous structures is uniform and consists of wurtzite‐ZnO throughout the film. Scanning electron microscopy reveals an average pore dimension of 30 nm. The potential of this material for a hybrid photovoltaic application is demonstrated by the manufacture of a poly(3‐hexylthiophene)/ZnO solar cell.     

Facile Fabrication of Monolithic 3D Porous Silica Microstructures and a Microfluidic System Embedded with the Microstructure

Monolithic 3D porous silica structures are fabricated into a multilayer framework with a bimodal pore size distribution in the micrometer and sub‐micrometer range. The fabrication – which involves directed assembly of colloidal spheres, transfer printing, and removal of a sacrificial template – yields robust and mechanically stable structures over a large area. The structure becomes monolithic upon pyrolyzing the stacked layers, which induces necking of the particles. The monolithic microstructures can easily be embedded in microchannels with the aid of photolithography, leading to the formation of a microfluidic system with a built‐in microstructure in a site‐ and shape‐controlled manner. Utilization of the system results in a fourfold increase in the mixing efficiency in the microchannel.     

Engineering of Mature Human Induced Pluripotent Stem Cell‐Derived Cardiomyocytes Using Substrates with Multiscale Topography

Producing mature and functional cardiomyocytes (CMs) by in vitro differentiation of induced pluripotent stem cells (iPSCs) using only biochemical cues is challenging. To mimic the biophysical and biomechanical complexity of the native in vivo environment during the differentiation and maturation process, polydimethylsiloxane substrates with 3D topography at the micrometer and sub‐micrometer levels are developed and used as cell‐culture substrates. The results show that while cylindrical patterns on the substrates resembling mature CMs enhance the maturation of iPSC‐derived CMs, sub‐micrometer‐level topographical features derived by imprinting primary human CMs further accelerate both the differentiation and maturation processes. The resulting CMs exhibit a more‐mature phenotype than control groups—as confirmed by quantitative polymerase chain reaction, flow cytometry, and the magnitude of beating signals—and possess the shape and orientation of mature CMs in human myocardium—as revealed by fluorescence microscopy, Ca²⁺ flow direction, and mitochondrial distribution. The experiments, combined with a virtual cell model, show that the physico‐mechanical cues generated by these 3D‐patterned substrates improve the phenotype of the CMs via the reorganization of the cytoskeletal network and the regulation of chromatin conformation.     

High‐Resolution Near‐Field Optical Investigation of Crystalline Domains in Oligomeric PQT‐12 Thin Films

The structure and morphology on different length scales dictate both the electrical and optical properties of organic semiconductor thin films. Using a combination of spectroscopic methods, including scanning near‐field optical microscopy, we study the domain structure and packing quality of highly crystalline thin films of oligomeric PQT‐12 with 100 nanometer spatial resolution. The pronounced optical anisotropy of these layers measured by polarized light microscopy facilitates the identification of regions with uniform molecular orientation. We find that a hierarchical order on three different length scales exists in these layers, made up of distinct well‐ordered dichroic areas at the ten‐micrometer‐scale, which are sub‐divided into domains with different molecular in‐plane orientation. These serve as a template for the formation of smaller needle‐like crystallites at the layer surface. A high degree of crystalline order is believed to be the cause of the rather high field‐effect mobility of these layers of 10^?3 cm² V^?1 s^?1, whereas it is limited by the presence of domain boundaries at macroscopic distances.     

Highly Anisotropic Suspended Planar‐Array Chips with Multidimensional Sub‐Micrometric Biomolecular Patterns

Suspended planar‐array (SPA) chips embody millions of individual miniaturized arrays to work in extremely small volumes. Here, the basis of a robust methodology for the fabrication of SPA silicon chips with on‐demand physical and chemical anisotropies is demonstrated. Specifically, physical traits are defined during the fabrication process with special focus on the aspect ratio, branching, faceting, and size gradient of the final chips. Additionally, the chemical attributes augment the functionality of the chips with the inclusion of complete coverage or patterns of selected biomolecules on the surface of the chips with contact printing techniques, offering an extremely high versatility, not only with the choice of the pattern shape and distribution but also in the choice of biomolecular inks to pattern. This approach increases the miniaturization of printed arrays in 3D structures by two orders of magnitude compared to those previously demonstrated. Finally, functional micrometric and sub‐micrometric patterned features are demonstrated with an antibody binding assay with the recognition of the printed spots with labeled antibodies from solution. The selective addition of physical and chemical attributes on the suspended chips represents the basis for future biomedical assays performed within extremely small volumes.     

Towards Sub‐Microscale Liquid Metal Patterns: Cascade Phase Change Mediated Pick‐n‐Place Transfer of Liquid Metals Printed and Stretched over a Flexible Substrate

Eutectic gallium indium (EGaIn) is actively investigated toward wearable and stretchable electronic devices due to the high fluidity, high electrical conductivity, and low toxicity. However, high surface tension along with spontaneous oxidation makes fine patterning below ≈10 µm challenging. In this paper, a novel manufacturing technique that enables EGaIn patterns of single‐digit micrometer widths on planar elastomeric substrates is presented. First, a custom direct printing setup is constructed for continuous and uniform printing of EGaIn by feedback control of the distance between the dispensing needle and the substrate. With this setup, a 120 µm wide linear pattern is printed on the Ecoflex, a stretchable elastomer. Then, the initial printed line is stretched, frozen with deionized water, and transferred to an unstretched Ecoflex substrate. Upon gentle heating after the pick‐n‐place of the EGaIn line frozen with deionized water, only the stretched EGaIn line is left on the new Ecoflex substrate. The aforementioned pick‐n‐place transfer of the stretched EGaIn frozen with water is cascaded multiple times until a target width is obtained. Finally, a 2 µm wide linear pattern, 60‐fold reduction with respect to the initial dimension, is acquired. For practical applications, strain and tactile sensors are demonstrated by width‐reduced EGaIn patterns.     

Three‐Dimensional Inkjet‐Printed Interconnects using Functional Metallic Nanoparticle Inks

Inkjet‐printed gold nanoparticle pillars are investigated as a high‐performance alternative to conventional flip‐chip interconnects for electronic packages, with significant advantages in terms of mechanical/chemical robustness and conductivity. The process parameters critical to pillar fabrication are described and highly uniform pillar arrays are demonstrated. More generally, this work underscores the impact of sintering on the electrical, mechanical, structural, and compositional properties of three‐dimensional nanoparticle‐based structures. Using heat treatments as low as 200 °C, electrical and mechanical performance that outcompetes conventional lead‐tin eutectic solder materials is achieved. With sintering conditions reaching 300 °C it is possible to achieve pillars with properties comparable to bulk gold. This work demonstrates the immense potential for both inkjet printing and metal nanoparticles to become a viable and cost‐saving alternative to both conventional electronic packaging processes and application‐specific integration schemes.     

Facile Fabrication of High‐Density Sub‐1‐nm Gaps from Au Nanoparticle Monolayers as Reproducible SERS Substrates

The fabrication of ultrasmall nanogaps (sub‐1 nm) with high density is of significant interest and importance in physics, chemistry, life science, materials science, surface science, nanotechnology, and environmental engineering. However, it remains a challenge to generate uncovered and clean sub‐1‐nm gaps with high density and uniform reproducibility. Here, a facile and low‐cost approach is demonstrated for the fabrication of high‐density sub‐1‐nm gaps from Au nanoparticle monolayers as reproducible surface‐enhanced Raman scattering (SERS) substrates. Au nanoparticles with larger diameters possess lower surface charge, thus the obtained large‐area nanoparticle monolayer generates a high‐density of sub‐1‐nm gaps. In addition, a remarkable SERS performance with a 10¹¹ magnitude for the Raman enhancement is achieved for 120 nm Au nanoparticle monolayers due to the dramatic increase in the electromagnetic field enhancement when the obtained gap is smaller than 0.5 nm. The Au nanoparticle monolayer is also transferred onto a stretchable PDMS substrate and the structural stability and reproducibility of the high‐density sub‐1‐nm gaps in Au monolayer films are illustrated. The resultant Au nanoparticle monolayer substrates with an increasing particle diameter exhibit tunable plasmonic properties, which control the plasmon‐enhanced photocatalytic efficiency for the dimerization of p‐aminothiophenol. The findings reported here offer a new opportunity for expanding the SERS application.     

Inside Front Cover: Inkjet Printing of Luminescent CdTe Nanocrystal–Polymer Composites (Adv. Funct. Mater. 1/2007)

Schubert and co‐workers have performed a detailed investigation on ink‐jet printing of well‐defined dots of luminescent CdTe nanocrystals (NCs) embedded in a poly(vinyl alcohol) matrix, as reported on p. 23, and subsequently made studies of their morphology and photoluminescence. The inside cover shows a photograph of an ink‐jet‐printed combinatorial library of differently sized CdTe NCs emitting at different wavelengths, and a 3D profilometer image of an array of printed dots. Inkjet printing is used to produce well‐defined patterns of dots (with diameters of ca. 120 μm) that are composed of luminescent CdTe nanocrystals (NCs) embedded within a poly(vinylalcohol) (PVA) matrix. Addition of ethylene glycol (1–2 vol %) to the aqueous solution of CdTe NCs suppresses the well‐known ring‐formation effect in inkjet printing leading to exceptionally uniform dots. Atomic force microscopy characterization reveals that in the CdTe NC films the particle–particle interaction could be prevented using inert PVA as a matrix. Combinatorial libraries of CdTe NC–PVA composites with variable NC sizes and polymer/NC ratios are prepared using inkjet printing. These libraries are subsequently characterized using a UV/fluorescence plate reader to determine their luminescent properties. Energy transfer from green‐light‐emitting to red‐light‐emitting CdTe NCs in the composite containing green‐ (2.6 nm diameter) and red‐emitting (3.5 nm diameter) NCs are demonstrated.     

Ultrasmall Au Nanoparticles Embedded in 2D Mixed‐Ligand Metal–Organic Framework Nanosheets Exhibiting Highly Efficient and Size‐Selective Catalysis

Metal–organic frameworks (MOFs) hold great promise as porous matrixes for the incorporation of Au nanoparticles (NPs) because of their rationally designed framework structures. Unfortunately, the as‐synthesized bulk MOFs usually vary in the range of micrometer or sub‐micrometer size, rendering extremely longer molecular diffusion distance of chemical species. 2D MOF nanosheets with extended lateral dimensions and nanometer thickness are expected to implement fast kinetics and effectively lower mass‐transfer barriers during embedding Au NPs process and sequential catalytic reactions. In this study, a novel 2D nanosheet of mixed‐ligand Ni(II) MOF (referred to NMOF‐Ni ) is successfully fabricated. With the merits of well‐defined micropores and functional oxygen‐decorated inner walls, the incorporation of quite monodisperse ultrasmall Au nanoparticles of around 1 nm into NMOF‐Ni has been achieved for the first time. The resulting nanocomposites exhibit remarkable catalytic performance and good size selectivity toward aqueous reduction reactions of nitrophenol, taking advantage of ultrasmall Au and 2D nanosheet nature, as well as the intact microporosity of host matrix. The present encouraging findings might shed light on new ways to develop high‐performance heterogeneous catalysts by using of 2D MOF nanosheets with functional cavities as hosts for homogeneous distribution of ultrasmall Au NPs.     

High‐Resolution Patterning of Hydrogels in Three Dimensions using Direct‐Write Photofabrication for Cell Guidance

The development of three‐dimensional, spatially defined neuronal cultures that mimic chemical and physical attributes of native tissue is of considerable interest for various applications, including the development of tailored neuronal networks and clinical repair of damaged nerves. Here, the use of multiphoton excitation to photocrosslink protein microstructures within three‐dimensional, optically transparent hydrogel materials, such as those based on hyaluronic acid, is reported. Multiphoton excitation confines photocrosslinking to a three‐dimensional voxel with submicron spatial resolution, enabling fabrication of protein matrices with low‐ to sub‐micrometer feature sizes by scanning the focus of a laser relative to the reagent solution. These methods can be used to create complex three‐dimensional architectures that provide both chemical and topographical cues for cell culture and guidance, providing for the first time a means to direct cell adhesion and migration on size scales relevant to in vivo environments. Using this approach, guidance of both dorsal root ganglion cells (DRGs) and hippocampal neural progenitor cells (NPCs) along arbitrary, three‐dimensional paths is demonstrated.     

Cover Picture: Fabrication and Characterization of Three‐Dimensional Silver‐Coated Polymeric Microstructures (Adv. Funct. Mater. 13/2006)

Kuebler and co‐workers report on p. 1739 a method for preparing conductive and optically reflective silver‐coated polymeric microstructures having virtually any 3D form. Shown are reflection images of a silvered five‐layer simple‐cubic lattice having a period of 2.4 μm (background) and a macroscopic silvered polymer film (inset). To prepare metallopolymeric microstructures, 3D polymeric scaffolds are first created by multiphoton direct laser writing, then functionalized with gold particles, and metallized using nucleated electroless silver deposition. A method is reported for fabricating complex 3D silver‐coated polymeric microstructures. The approach is based on the creation of a crosslinked polymeric microscaffold via patterned multiphoton‐initiated polymerization followed by surface‐nucleated electroless deposition of silver. The conductivity and reflectivity of the resulting silver–polymer composites and the nanoscale morphology of the deposited silver are characterized. Sub‐micrometer thick layers of silver can be controllably deposited onto surfaces, including those of 3D microporous forms without occluding the interior of the structure. The approach is general for silver coating crosslinked polymeric structures based on acrylate, methacrylate, and epoxide resins and provides a new path to complex 3D micrometer‐scale devices with electronic, photonic, and electromechanical function.     

Dynamic Photomask‐Assisted Direct Ink Writing Multimaterial for Multilevel Triboelectric Nanogenerator

Triboelectric nanogenerator (TENG) devices are extensively studied as a mechanical energy harvester and self‐powered sensor for wearable electronics and physiological monitoring. However, the conventional TENG fabrication involving assembling steps and using the single property of matrix material suffers from simple devices shape and a single level of mechanical response for sensing and energy harvesting. Here, the printed multimaterial matrix for multilevel mechanical‐responsive TENG with on‐demand reconfiguration of shape is reported. A multimaterial 3D printing approach by using dynamic photomask‐assisted direct ink writing printing together with a two‐stage curing hybrid ink is first developed. Multimaterial structures with location‐specific properties, such as tensile modulus, failure stress, and glass transition temperature for controlled deformation, crack propagation path, and sequential shape memory, are directly printed. The printed multimaterial structure with sequential deformation behavior is used to fabricate a multilevel‐TENG (mTENG) device for multiple level mechanical energy harvesters and sensors. It is demonstrated that the mTENG can be embedded in shoe insoles to achieve both comfortable wearing and motion state monitoring. This work provides a new approach to combine multimaterial 3D printing with TENG devices for functional wearable electronics as energy harvester and sensors.     

Helical 3D‐Printed Metal Electrodes as Custom‐Shaped 3D Platform for Electrochemical Devices

3D‐printing represents an emerging technology that can revolutionize the way object and functional devices are fabricated. Here the use of metal 3D printing is demonstrated to fabricate bespoke electrochemical stainless steel electrodes that can be used as platform for different electrochemical applications ranging from electrochemical capacitors, oxygen evolution catalyst, and pH sensor by means of an effective and controlled deposition of IrO₂ films. The electrodes have been characterized by scanning electrode microscopy and energy dispersive X‐ray spectroscopy before the electrochemical testing. Excellent pseudocapacitive as well as catalytic properties have been achieved with these 3D printed steel‐IrO₂ electrodes in alkaline solutions. These electrodes also demonstrate Nernstian behavior as pH sensor. This work represents a breakthrough in on‐site prototyping and fabrication of highly tailored electrochemical devices with complex 3D shapes which facilitate specific functions and properties.     

Inkjet Printing of Luminescent CdTe Nanocrystal–Polymer Composites

Inkjet printing is used to produce well‐defined patterns of dots (with diameters of ca. 120 μm) that are composed of luminescent CdTe nanocrystals (NCs) embedded within a poly(vinylalcohol) (PVA) matrix. Addition of ethylene glycol (1–2 vol %) to the aqueous solution of CdTe NCs suppresses the well‐known ring‐formation effect in inkjet printing leading to exceptionally uniform dots. Atomic force microscopy characterization reveals that in the CdTe NC films the particle–particle interaction could be prevented using inert PVA as a matrix. Combinatorial libraries of CdTe NC–PVA composites with variable NC sizes and polymer/NC ratios are prepared using inkjet printing. These libraries are subsequently characterized using a UV/fluorescence plate reader to determine their luminescent properties. Energy transfer from green‐light‐emitting to red‐light‐emitting CdTe NCs in the composite containing green‐ (2.6 nm diameter) and red‐emitting (3.5 nm diameter) NCs are demonstrated.     

Light‐Controlled,High‐Resolution Patterning of Living Engineered Bacteria Onto Textiles,Ceramics, and Plastic

Living cells can impart materials with advanced functions, such as sense‐and‐respond, chemical production, toxin remediation, energy generation and storage, self‐destruction, and self‐healing. Here, an approach is presented to use light to pattern Escherichia coli onto diverse materials by controlling the expression of curli fibers that anchor the formation of a biofilm. Different colors of light are used to express variants of the structural protein CsgA fused to different peptide tags. By projecting color images onto the material containing bacteria, this system can be used to pattern the growth of composite materials, including layers of protein and gold nanoparticles. This is used to pattern cells onto materials used for 3D printing, plastics (polystyrene), and textiles (cotton). Further, the adhered cells are demonstrated to respond to sensory information, including small molecules (IPTG and DAPG) and light from light‐emitting diodes. This work advances the capacity to engineer responsive living materials in which cells provide diverse functionality.     

Nanogold‐Loaded Sharp‐Edged Carbon Bullets as Plant‐Gene Carriers

The higher DNA delivery efficiency into plants by gold nanoparticles embedded in sharp carbonaceous carriers is demonstrated. These nanogold‐embedded carbon matrices are prepared by heat treatment of biogenic intracellular gold nanoparticles. The DNA‐delivery efficiency is tested on a model plant, Nicotiana tabacum, and is further extended to the monocot, Oryza sativa, and a hard dicot tree species, Leucaena leucocephala. These materials reveal good dispersion of the transport material, producing a greater number of GUS foci per unit area. The added advantages of the composite carrier are the lower plasmid and gold requirements. Plant‐cell damage with the carbon‐supported particles is very minimal and can be gauged from the increased plant regeneration and transformation efficiency compared with that of the commercial micrometer‐sized gold particles. This is ascribed to the sharp edges that the carbon supports possess, which lead to better piercing capabilities with minimum damage.     

High‐Density Hotspots Engineered by Naturally Piled‐Up Subwavelength Structures in Three‐Dimensional Copper Butterfly Wing Scales for Surface‐Enhanced Raman Scattering Detection

Very recently, wing scales of natural Lepidopterans (butterflies and moths) manifested themselves in providing excellent three dimensional (3D) hierarchical structures for surface‐enhanced Raman scattering (SERS) detection. But the origin of the observed enormous Raman enhancement of the analytes on 3D metallic replicas of butterfly wing scales has not been clarified yet, hindering a full utilization of this huge natural wealth with more than 175 000 3D morphologies. Herein, the 3D sub‐micrometer Cu structures replicated from butterfly wing scales are successfully tuned by modifying the Cu deposition time. An optimized Cu plating process (10 min in Cu deposition) yields replicas with the best conformal morphologies of original wing scales and in turn the best SERS performance. Simulation results show that the so‐called “rib‐structures” in Cu butterfly wing scales present naturally piled‐up hotspots where electromagnetic fields are substantially amplified, giving rise to a much higher hotspot density than in plain 2D Cu structures. Such a mechanism is further verified in several Cu replicas of scales from various butterfly species. This finding paves the way to the optimal scale candidates out of ca. 175 000 Lepidopteran species as bio‐templates to replicate for SERS applications, and thus helps bring affordable SERS substrates as consumables with high sensitivity, high reproducibility, and low cost to ordinary laboratories across the world.