Quantitative nanopatterning of fg-scale liquids with dip-pen nanolithography.

The ability to precisely pattern nanoscale amounts of liquids is essential for biotechnology and high-throughput chemistry, but controlling fluid flow on these scales is very challenging. Scanning probe lithography methods such as dip-pen nanolithography (DPN) provide a mechanism to write fluids at the nanoscale, but this is an open loop process as methods to provide feedback while patterning sub-pg features have yet to be reported. Here, we demonstrate a novel method for programmably nanopatterning liquid features at the fg-scale through a combination of ultrafast atomic force microscopy probes, the use of spherical tips, and inertial mass sensing. We begin by investigating the required probe properties that would provide sufficient mass responsivity to detect fg-scale mass changes and find ultrafast probes to be capable of this resolution. Further, we attach a spherical bead to the tip of an ultrafast probe as we hypothesize that the spherical tip could hold a drop at its apex which both facilitates interpretation of inertial sensing and maintains a consistent fluid environment for reliable patterning. We experimentally find that sphere-tipped ultrafast probes are capable of reliably patterning hundreds of features in a single experiment. Analyzing the changes in the vibrational resonance frequency during the patterning process, we find that drift in the resonance frequency complicates analysis, but that it can be removed through a systematic correction. Subsequently, we quantitatively study patterning using sphere-tipped ultrafast probes as a function of retraction speed and dwell time to find that the mass of fluid transferred can be modulated by greater than an order of magnitude and that liquid features as small as 6 fg can be patterned and resolved. Taken together, this work addresses a persistent concern in DPN by enabling quantitative feedback for nanopatterning of aL-scale features and lays the foundation for programmably nanopatterning fluids.

Patterning of Hydrophilic and Hydrophobic Gold and Magnetite Nanoparticles by Dip Pen Nanolithography.

Nanoparticles offer unique physical and chemical properties. Dip pen nanolithography of nanoparticles enables versatile patterning and nanofabrication with potential application in electronics and sensing, but is not well studied yet. Herein, the patterned deposition of various nanoparticles onto unmodified silicon substrates is presented. It is shown that aqueous solutions of hydrophilic citrate and cyclodextrin functionalized gold nanoparticles as well as poly(acrylic) acid decorated magnetite nanoparticles are feasible for writing nanostructures. Both smaller and larger nanoparticles can be patterned. Hydrophobic oleylamine or n-dodecylamine capped gold nanoparticles and oleic acid decorated magnetite nanoparticles are deposited from toluene. Tip loading is carried out by dip-coating, and writing succeeds fast within 0.1 s. Also, coating with longer tip dwell times, at different relative humidity and varying frequency are studied for deposition of nanoparticle clusters. The resulting feature size is between 300 and 1780 nm as determined by scanning electron microscopy. Atomic force microscopy confirms that the heights of the deposited structures correspond to a single or double layer of nanoparticles. Higher writing speeds lead to smaller line thicknesses, offering possibilities to more complex structures. Dip pen nanolithography can hence be used to pattern nanoparticles on silicon substrates independent of the surface chemistry.

3D Nanolithography by Means of Lipid Ink Spreading Inhibition.

While patterning 2D metallic nanostructures are well established through different techniques, 3D printing still constitutes a major bottleneck on the way to device miniaturization. In this work a fluid phase phospholipid ink is used as a building block for structuring with dip-pen nanolithography. Following a bioinspired approach that relies on ink-spreading inhibition, two processes are presented to build 2D and 3D metallic structures. Serum albumin, a widely used protein with an innate capability to bind to lipids, is the key in both processes. Covering the sample surface with it prior to lipid writing, anchors lipids on the substrate, which ultimately allows the creation of highly stable 3D lipid-based scaffolds to build metallic structures.

Dip-Pen Nanolithography(DPN): from Micro/Nano-patterns to Biosensing.

Dip-pen nanolithography is an emerging and attractive surface modification technique that has the capacity to directly and controllably write micro/nano-array patterns on diverse substrates. The superior throughput, resolution, and registration enable DPN an outstanding candidate for biological detection from the molecular level to the cellular level. Herein, we overview the technological evolution of DPN in terms of its advanced derivatives and DPN-enabled versatile sensing patterns featuring multiple compositions and structures for biosensing. Benefitting from uniform, reproducible, and large-area array patterns, DPN-based biosensors have shown high sensitivity, excellent selectivity, and fast response in target analyte detection and specific cellular recognition. We anticipate that DPN-based technologies could offer great potential opportunities to fabricate multiplexed, programmable, and commercial array-based sensing biochips.

Rapid Capture of Cancer Extracellular Vesicles by Lipid Patch Microarrays.

Extracellular vesicles (EVs) contain various bioactive molecules such as DNA, RNA, and proteins, and play a key role in the regulation of cancer progression. Furthermore, cancer-associated EVs carry specific biomarkers and can be used in liquid biopsy for cancer detection. However, it is still technically challenging and time consuming to detect or isolate cancer-associated EVs from complex biofluids (e.g., blood). Here, a novel EV-capture strategy based on dip-pen nanolithography generated microarrays of supported lipid membranes is presented. These arrays carry specific antibodies recognizing EV- and cancer-specific surface biomarkers, enabling highly selective and efficient capture. Importantly, it is shown that the nucleic acid cargo of captured EVs is retained on the lipid array, providing the potential for downstream analysis. Finally, the feasibility of EV capture from patient sera is demonstrated. The demonstrated platform offers rapid capture, high specificity, and sensitivity, with only a small need in analyte volume and without additional purification steps. The platform is applied in context of cancer-associated EVs, but it can easily be adapted to other diagnostic EV targets by use of corresponding antibodies.

Closed-Loop Nanopatterning of Liquids with Dip-Pen Nanolithography.

The ability to reliably manipulate small quantities of liquids is the backbone of high-throughput chemistry, but the continual drive for miniaturization necessitates creativity in how nanoscale samples of liquids are handled. Here, we describe a closed-loop method for patterning liquid samples on pL to sub-fL scales using scanning probe lithography. Specifically, we employ tipless scanning probes and identify liquid properties that enable probe-sample transport that is readily tuned using probe withdrawal speed. Subsequently, we introduce a novel two-harmonic inertial sensing scheme for tracking the mass of liquid on the probe. Finally, this is combined with a fluid mechanics-based iterative control scheme that selects printing conditions to meet a target feature mass to enable closed-loop patterning with better than 1% accuracy and ∼4% precision in terms of mass. Taken together, these advances address a pervasive issue in scanning probe lithography, namely, real-time closed-loop control over patterning, and position scanning probe lithography of liquids as a candidate for the robust nanoscale manipulation of liquids for advanced high-throughput chemistry.

Controlling the Size and Pattern Pitch of Ni(OH)2 Nanoclusters Using Dip-Pen Nanolithography to Improve Water Oxidation.

We use dip-pen nanolithography to accurately pattern Ni(OH)2 nanoclusters on a metachemical surface with an exceptionally large surface area. The distance between the nanoclusters can be manipulated to control the oxygen-evolution reaction current and overpotential, thereby improving the efficiency of the water-splitting process while using minute amounts of the catalyst.

Enhanced Stability of Lipid Structures by Dip-Pen Nanolithography on Block-Type MPC Copolymer.

Biomimetic lipid membranes on solid supports have been used in a plethora of applications, including as biosensors, in research on membrane proteins or as interfaces in cell experiments. For many of these applications, structured lipid membranes, e.g., in the form of arrays with features of different functionality, are highly desired. The stability of these features on a given substrate during storage and in incubation steps is key, while at the same time the substrate ideally should also exhibit antifouling properties. Here, we describe the highly beneficial properties of a 2-methacryloyloxyethyl phosphorylcholine (MPC) copolymer for the stability of supported lipid membrane structures generated by dip-pen nanolithography with phospholipids (L-DPN). The MPC copolymer substrates allow for more stable and higher membrane stack structures in comparison to other hydrophilic substrates, like glass or silicon oxide surfaces. The structures remain highly stable under immersion in liquid and subsequent incubation and washing steps. This allows multiplexed functionalization of lipid arrays with antibodies via microchannel cantilever spotting (µCS), without the need of orthogonal binding tags for each antibody type. The combined properties of the MPC copolymer substrate demonstrate a great potential for lipid-based biomedical sensing and diagnostic platforms.

Enhanced Molecular Spin-Photon Coupling at Superconducting Nanoconstrictions.

We combine top-down and bottom-up nanolithography to optimize the coupling of small molecular spin ensembles to 1.4 GHz on-chip superconducting resonators. Nanoscopic constrictions, fabricated with a focused ion beam at the central transmission line, locally concentrate the microwave magnetic field. Drops of free-radical molecules have been deposited from solution onto the circuits. For the smallest ones, the molecules were delivered at the relevant circuit areas by means of an atomic force microscope. The number of spins Neff effectively coupled to each device was accurately determined combining Scanning Electron and Atomic Force Microscopies. The collective spin-photon coupling constant has been determined for samples with Neff ranging between 2 × 106 and 1012 spins, and for temperatures down to 44 mK. The results show the well-known collective enhancement of the coupling proportional to the square root of Neff. The average coupling of individual spins is enhanced by more than 4 orders of magnitude (from 4 mHz up to above 180 Hz), when the transmission line width is reduced from 400 µm down to 42 nm, and reaches maximum values near 1 kHz for molecules located on the smallest nanoconstrictions.

Scanning Probe-Directed Assembly and Rapid Chemical Writing Using Nanoscopic Flow of Phospholipids.

Nanofluidic systems offer a huge potential for discovery of new molecular transport and chemical phenomena that can be employed for future technologies. Herein, we report on the transport behavior of surface-reactive compounds in a nanometer-scale flow of phospholipids from a scanning probe. We have investigated microscopic deposit formation on polycrystalline gold by lithographic printing and writing of 1,2-dioleoyl-sn-glycero-3-phosphocholine and eicosanethiol mixtures, with the latter compound being a model case for self-assembled monolayers (SAMs). By analyzing the ink transport rates, we found that the transfer of thiols was fully controlled by the fluid lipid matrix allowing to achieve a certain jetting regime, i.e., transport rates previously not reported in dip-pen nanolithography (DPN) studies on surface-reactive, SAM-forming molecules. Such a transport behavior deviated significantly from the so-called molecular diffusion models, and it was most obvious at the high writing speeds, close to 100 µm s-1. Moreover, the combined data from imaging ellipsometry, scanning electron microscopy, atomic force microscopy (AFM), and spectroscopy revealed a rapid and efficient ink phase separation occurring in the AFM tip-gold contact zone. The force curve analysis indicated formation of a mixed ink meniscus behaving as a self-organizing liquid. Based on our data, it has to be considered as one of the co-acting mechanisms driving the surface reactions and self-assembly under such highly nonequilibrium, crowded environment conditions. The results of the present study significantly extend the capabilities of DPN using standard AFM instrumentation: in the writing regime, the patterning speed was already comparable to that achievable by using electron beam systems. We demonstrate that lipid flow-controlled chemical patterning process is directly applicable for rapid prototyping of solid-state devices having mesoscopic features as well as for biomolecular architectures.

Development of Dip-Pen Nanolithography (DPN) and Its Derivatives.

Dip-pen nanolithography (DPN) is a unique nanofabrication tool that can directly write a variety of molecular patterns on a surface with high resolution and excellent registration. Over the past 20 years, DPN has experienced a tremendous evolution in terms of applicable inks, a remarkable improvement in fabrication throughput, and the development of various derivative technologies. Among these developments, polymer pen lithography (PPL) is the most prominent one that provides a large-scale, high-throughput, low-cost tool for nanofabrication, which significantly extends DPN and beyond. These developments not only expand the scope of the wide field of scanning probe lithography, but also enable DPN and PPL as general approaches for the fabrication or study of nanostructures and nanomaterials. In this review, a focused summary and historical perspective of the technological development of DPN and its derivatives, with a focus on PPL, in one timeline, are provided and future opportunities for technological exploration in this field are proposed.

Locally Controlled Growth of Individual Lambda-Shaped Carbon Nanofibers.

The locally defined growth of carbon nanofibers with lambda shape in an open flame process is demonstrated. Via the growth time, the geometry of the structures can be tailored to a Λ- or λ-type shape. Microchannel cantilever spotting and dip-pen nanolithography are utilized for the deposition of catalytic salt NiCl2 · 6H2 O for locally controlled growth of lambda-shaped carbon nanofibers. Rigorous downscaling reveals a critical catalytic salt volume of 0.033 µm³, resulting in exactly one lambda-shaped carbon nanofiber at a highly predefined position. An empirical model explains the observed growth process.

Multiplexed Biomolecular Arrays Generated via Parallel Dip-Pen Nanolithography.

The capability of transferring target materials especially functionality-reliable biomolecules, into specific locations and with arbitrarily designed patterns are of critical importance for high-throughput disease diagnosis, multiplexing, and drug screening. Herein, we report the simultaneous patterning of two types of biomolecules using the parallel dip-pen nanolithography technology where an array of the atomic force microscope (AFM) tips can be selectively and alternately coated with target biomolecules via a specially designed inkwell array. Moreover, mixing target biomolecules at a proper volumetric ratio with polyethylene glycol dissolved in PBS buffer solution that works as an ink carrier can not only facilitate the smooth transfer of ink materials from the AFM tip to the substrate, it can also help to adjust the ink diffusion constant of different biomolecules to be highly similar so that the multiplexed biofunctional dot and/or line arrays at similar sizes can be reliably generated.

The Role of Liquid Ink Transport in the Direct Placement of Quantum Dot Emitters onto Sub-Micrometer Antennas by Dip-Pen Nanolithography.

Dip-pen nanolithography (DPN) is used to precisely position core/thick-shell ("giant") quantum dots (gQDs; ≥10 nm in diameter) exclusively on top of silicon nanodisk antennas (≈500 nm diameter pillars with a height of ≈200 nm), resulting in periodic arrays of hybrid nanostructures and demonstrating a facile integration strategy toward next-generation quantum light sources. A three-step reading-inking-writing approach is employed, where atomic force microscopy (AFM) images of the pre-patterned substrate topography are used as maps to direct accurate placement of nanocrystals. The DPN "ink" comprises gQDs suspended in a non-aqueous carrier solvent, o-dichlorobenzene. Systematic analyses of factors influencing deposition rate for this non-conventional DPN ink are described for flat substrates and used to establish the conditions required to achieve small (sub-500 nm) feature sizes, namely: dwell time, ink-substrate contact angle and ink volume. Finally, it is shown that the rate of solvent transport controls the feature size in which gQDs are found on the substrate, but also that the number and consistency of nanocrystals deposited depends on the stability of the gQD suspension. Overall, the results lay the groundwork for expanded use of nanocrystal liquid inks and DPN for fabrication of multi-component nanostructures that are challenging to create using traditional lithographic techniques.

Direct-Patterning SWCNTs Using Dip Pen Nanolithography for SWCNT/Silicon Solar Cells.

Dip pen nanolithography (DPN) is used to pattern single-walled carbon nanotube (SWCNT) lines between the n-type Si and SWCNT film in SWCNT/Si solar cells. The SWCNT ink composition, loading, and DPN pretreatment are optimized to improve patterning. This improved DPN technique is then used to successfully pattern >1 mm long SWCNT lines consistently. This is a 20-fold increase in the previously reported direct-patterning of SWCNT lines using the DPN technique, and demonstrates the scalability of the technique to pattern larger areas. The degree of the uniformity of SWCNTs in these lines is further characterized by Raman spectroscopy and scanning electron microscopy. The patterned SWCNT lines are used as thin conductive pathways in SWCNT/Si solar cells, similar to front contact electrodes. The critical parameters of these solar cells are measured and compared to control cells without SWCNT lines. The addition of SWCNT lines increases power conversion efficiency by 40% (relative). Importantly, the SWCNT lines reduce average series resistance by 44%, and consequently increase average fill factor by 24%.

Molecular Patterning and Directed Self-Assembly of Gold Nanoparticles on GaAs.

The ability to create micro-/nanopatterns of organic self-assembled monolayers (SAMs) on semiconductor surfaces is crucial for fundamental studies and applications in a number of emerging fields in nanoscience. Here, we demonstrate the direct patterning of thiolate SAMs on oxide-free GaAs surface by dip-pen nanolithography (DPN) and microcontact printing (µCP), facilitated by a process of surface etching and passivation of the GaAs. A quantitative analysis on the molecular diffusion on GaAs was conducted by examining the writing of nanoscale dot and line patterns by DPN, which agrees well with surface diffusion models. The functionality of the patterned thiol molecules was demonstrated by directed self-assembly of gold nanoparticles (Au NPs) onto a template of 4-aminothiophenol (ATP) SAM on GaAs. The highly selective assembly of the Au NPs was made evident with atomic force microscopy (AFM) and scanning electron microscopy (SEM). The ability to precisely control the assembly of Au NPs on oxide-free semiconductor surfaces using molecular templates may lead to an efficient bottom-up method for the fabrication of nanoplasmonic structures.

Biomimetic Phospholipid Membrane Organization on Graphene and Graphene Oxide Surfaces: A Molecular Dynamics Simulation Study.

Supported phospholipid membrane patches stabilized on graphene surfaces have shown potential in sensor device functionalization, including biosensors and biocatalysis. Lipid dip-pen nanolithography (L-DPN) is a method useful in generating supported membrane structures that maintain lipid functionality, such as exhibiting specific interactions with protein molecules. Here, we have integrated L-DPN, atomic force microscopy, and coarse-grained molecular dynamics simulation methods to characterize the molecular properties of supported lipid membranes (SLMs) on graphene and graphene oxide supports. We observed substantial differences in the topologies of the stabilized lipid structures depending on the nature of the surface (polar graphene oxide vs nonpolar graphene). Furthermore, the addition of water to SLM systems resulted in large-scale reorganization of the lipid structures, with measurable effects on lipid lateral mobility within the supported membranes. We also observed reduced lipid ordering within the supported structures relative to free-standing lipid bilayers, attributed to the strong hydrophobic interactions between the lipids and support. Together, our results provide insight into the molecular effects of graphene and graphene oxide surfaces on lipid bilayer membranes. This will be important in the design of these surfaces for applications such as biosensor devices.

Attoliter Chemistry for Nanoscale Functionalization of Graphene.

The nanoscale, multiplexed functionalization of graphene in a device array is a critical step to realize graphene-based chemical and biosensors. We demonstrate that graphene can be functionalized with submicron resolution and in well-defined locations and patterns using reaction agents in attoliter quantities, utilizing dip-pen nanolithography or microchannel cantilever spotting. Specifically, we functionalize graphene with a biotin azide using click-chemistry and demonstrate the subsequent binding of fluorescently tagged streptavidin. The technique can be scaled up to multiplex functionalize graphene devices on a wafer-scale for sensor and biomedical applications.

Thermoresponsive Polymer Micropatterns Fabricated by Dip-Pen Nanolithography for a Highly Controllable Substrate with Potential Cellular Applications.

We report a novel approach for patterning thermoresponsive hydrogels based on N,N-diethylacrylamide (DEAAm) and bifunctional Jeffamine ED-600 by dip-pen nanolithography (DPN). The direct writing of micron-sized thermoresponsive polymer spots was achieved with efficient control over feature size. A Jeffamine-based ink prepared through the combination of organic polymers, such as DEAAm, in an inorganic silica network was used to print thermosensitive arrays on a thiol-silanized silicon oxide substrate. The use of a Jeffamine hydrogel, acting as a carrier matrix, allowed a reduction in the evaporation of ink molecules with high volatility, such as DEAAm, and facilitated the transfer of ink from tip to substrate. The thermoresponsive behavior of polymer arrays which swell/deswell in aqueous solution in response to a change in temperature was successfully characterized by atomic force microscopy (AFM) and Raman spectroscopy: a thermally induced change in height and hydration state was observed, respectively. Finally, we demonstrate that cells can adhere to and interact with these dynamic features and exhibit a change in behavior when cultured on the substrates above and below the transition temperature of the Jeffamine/DEAAm thermoresponsive hydrogels. This demonstrates the potential of these micropatterned hydrogels to act as a controllable surface for cell growth.

Biomimicking Nano-Micro Binary Polymer Brushes for Smart Cell Orientation and Adhesion Control.

A new biomimetic surface named nano-micro binary polymer brushes is fabricated by large-area bench-top dip-pen nanodisplacement lithography technique. It is composed of gelatin-modified poly(glycidyl methacrylate) nanolines which are spaced by microstripes of poly(N-isopropylacrylamide). Cells are not only adhered and oriented well on the re-used surface, but also detachable from the surface with well-preserved extracellular matrix and aligned morphology.