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Tagging, tracking, or validation of products are often facilitated by inkjet-printed optical information labels. However, this requires thorough substrate pretreatment, ink optimization, and often lacks in printing precision/resolution. Herein, a printing method based on laser-driven deposition of solid polymer ink that allows for printing on various substrates without pretreatment is demonstrated. Since the deposition process has a precision of

The identification of specific biomarkers for Zika infection and its clinical complications is fundamental to mitigate the infection spread, which has been associated with a broad range of neurological sequelae. We present the characterization of antibody responses in serum samples from individuals infected with Zika, presenting non-severe (classical) and severe (neurological disease) phenotypes, with high-density peptide arrays comprising the Zika NS1 and NS2B proteins. The data pinpoints one strongly IgG-targeted NS2B epitope in non-severe infections, which is absent in Zika patients, where infection progressed to the severe phenotype. This differential IgG profile between the studied groups was confirmed by multivariate data analysis. Molecular dynamics simulations and circular dichroism have shown that the peptide in solution presents itself in a sub-optimal conformation for antibody recognition, which led us to computationally engineer an artificial protein able to stabilize the NS2B epitope structure. The engineered protein was used to interrogate paired samples from mothers and their babies presenting Zika-associated microcephaly and confirmed the absence of NS2B IgG response in those samples. These findings suggest that the assessment of antibody responses to the herein identified NS2B epitope is a strong candidate biomarker for the diagnosis and prognosis of Zika-associated neurological disease.

A relative humidity sensor was produced by carbon laser patterning of a carbon precursor ink on a flexible substrate. Citric acid and urea, both inexpensive and naturally abundant molecules, are used as initial precursors to obtain a porous carbon foam after CO2laser irradiation. The laser-patterned material is characterized by electron microscopy, Raman spectroscopy, and vertical scanning interferometry. An intrinsic p-type semiconducting behavior was confirmed by thermoelectric and Hall measurements. The resistance of this porous, metal-free material is sensitive to atmospheric variations, namely, temperature and relative humidity (≈5Ω·%). Under dry atmosphere, the sensor acts as a thermometer with a linear relationship between temperature and relative variation of resistance (0.07%·K−1). The evolution of the sensor resistance at different relative humidities and temperatures is studied by electrical impedance measurements. The kinetic transitory regime of water desorption from the carbonaceous surface of the sensor is analyzed using Langmuir’s model. The equilibrium constant of adsorption Kads has been determined, and the standard enthalpy of adsorption of water on the sensor surface is estimated atΔadsH°=−42.6 kJ·mol−1. The simple and inexpensive production and its high, stable sensitivity make laser-patterned carbon interesting for humidity sensing applications, and the method allows for the large-scale production of printed sensor arrays.

Polymer modification plays an important role in the construction of devices, but the lack of fundamental understanding on polymer-surface adhesion limits the development of miniaturized devices. In this work, a thermoplastic polymer collection was established using the combinatorial laser-induced forward transfer technique as a research platform, to assess the adhesion of polymers to substrates of different wettability. Furthermore, it also revealed the influence of adhesion on dewetting phenomena during the laser transfer and relaxation process, resulting in polymer spots of various morphologies. This gives a general insight into polymer-surface adhesion and connects it with the generation of defined polymer microstructures, which can be a valuable reference for the rational use of polymers.

Laser-induced forward transfer (LIFT) is a rapid laser-patterning technique for high-throughput combinatorial synthesis directly on glass slides. A lack of automation and precision limited LIFT applications to simple proof-of-concept syntheses of fewer than 100 compounds. Here, we report an automated synthesis instrument that combines laser transfer and robotics for parallel synthesis in a microarray format with up to 10000 individual reactions/cm2. An optimized pipeline for amide bond formation is the basis for preparing complex peptide microarrays with thousands of different sequences in high yield with high reproducibility. The resulting peptide arrays are of higher quality than commercial peptide arrays. More than 4800 15-residue peptides resembling the entire Ebola virus proteome on a microarray were synthesized to study the antibody response of an Ebola virus infection survivor. We identified known and unknown epitopes that serve now as a basis for Ebola diagnostic development. The versatility and precision of the synthesizer is demonstrated by in situ synthesis of fluorescent molecules via Schiff base reaction and multi-step patterning of precisely definable amounts of fluorophores. This automated laser transfer synthesis approach opens new avenues for high-throughput chemical synthesis and biological screening.

Laser-induced forward transfer (LIFT) has the potential to be an alternative approach to atomic force microscopy based scanning probe lithography techniques, which have limitations in high-speed and large-scale patterning. However, traditional donor slides limit the resolution and chemical flexibility of LIFT. Here, we propose a hematite nanolayer absorber for donor slides to achieve high-resolution transfers down to sub-femtoliters. Being wettable by both aqueous and organic solvents, this new donor significantly increases the chemical scope for the LIFT process. For parallel amino acid coupling reactions, the patterning resolution can now be increased more than five times (>111,000 spots/cm2 for hematite donor versus 20,000 spots/cm2 for standard polyimide donor) with even faster scanning (2 versus 6 ms per spot). Due to the increased chemical flexibility, we could explore other types of reactions inside ultrasmall polymer reactors: copper (I) catalyzed click chemistry and laser-driven oxidation of a tetrahydroisoquinoline derivative, suggesting the potential of LIFT for both deposition of chemicals and laser-driven photochemical synthesis in femtoliters within milliseconds. Since the hematite shows no damage after typical laser transfer, donors can be regenerated by heat treatment. These findings will transform the LIFT process into an automatable, precise, and highly efficient technology for high-throughput femtoliter chemistry.

Automated chemical synthesis has revolutionized synthetic access to biopolymers in terms of simplicity and speed. While automated oligosaccharide synthesis has become faster and more versatile, the parallel synthesis of oligosaccharides is not yet possible. Here, a chemical vapor glycosylation strategy (VaporSPOT) is described that enables the simultaneous synthesis of oligosaccharides on a cellulose membrane solid support. Different linkers allow for flexible and straightforward cleavage, purification, and characterization of the target oligosaccharides. This method is the basis for the development of parallel automated glycan synthesis platforms.

We report the automated glycan assembly (AGA) of different oligosaccharide fragments of the bacterial peptidoglycan (PGN) backbone. Iterative addition on a solid support of an acetyl glucosamine and a new muramic acid building block is followed by cleavage from the solid support and final deprotection providing 10 oligosaccharides up to six units.

Considerable research efforts have been devoted to surface‐enhanced Raman spectroscopy (SERS), due to its excellent performance in biosensing and imaging. Here, a novel and facile strategy for the fabrication of well‐defined and uniform nanodimers as SERS substrates is presented. By the assistance of ultrasound, the violent polyol process for particle generation becomes controllable, enabling the self‐assembly of nanostars to nanodimers. Moreover, the aggregation of nanodimers can be easily tuned by post‐ultrasonic treatment, which gives a sensitive substrate for SERS., Made of nanostars: By the assistance of ultrasound, the violent polyol process for silver particle generation becomes controllable, enabling the self‐assembly of nanostars to nanodimers. Moreover, the aggregation of nanodimers can be easily tuned by post‐ultrasonic treatment, which gives a sensitive substrate for SERS.

High-density peptide arrays are an excellent means to profile anti-plasmodial antibody responses. Different protein intrinsic epitopes can be distinguished, and additional insights are gained, when compared with assays involving the full-length protein. Distinct reactivities to specific epitopes within one protein may explain differences in published results, regarding immunity or susceptibility to malaria. We pursued three approaches to find specific epitopes within important plasmodial proteins, (1) twelve leading vaccine candidates were mapped as overlapping 15-mer peptides, (2) a bioinformatical approach served to predict immunogenic malaria epitopes which were subsequently validated in the assay, and (3) randomly selected peptides from the malaria proteome were screened as a control. Several peptide array replicas were prepared, employing particle-based laser printing, and were used to screen 27 serum samples from a malaria-endemic area in Burkina Faso, West Africa. The immunological status of the individuals was classified as “protected” or “unprotected” based on clinical symptoms, parasite density, and age. The vaccine candidate screening approach resulted in significant hits in all twelve proteins and allowed us (1) to verify many known immunogenic structures, (2) to map B-cell epitopes across the entire sequence of each antigen and (3) to uncover novel immunogenic epitopes. Predicting immunogenic regions in the proteome of the human malaria parasite Plasmodium falciparum, via the bioinformatics approach and subsequent array screening, confirmed known immunogenic sequences, such as in the leading malaria vaccine candidate CSP and discovered immunogenic epitopes derived from hypothetical or unknown proteins.

Fluorescence signals have been widely used in information encryption for a few decades, but still suffer from limited reliability. Here, reversible multichannel fluorescent devices with encrypted information were constructed, based on two fluorescent positional isomers of a diphenylquinoxaline derivative. Possessing the same core fluorescent group and acid-/pH-responsive mechanism, the two isomers showed different fluorescence colors in an acidic environment; this allowed us to realize stepwise encryption of information in orthogonal fluorescence channels. Because the protonation was reversible, the revealed information could be re-encrypted simply by heating. This approach highlights the value of positional isomers to build multichannel encryption devices, improving their reliability on the molecular level.

Fabrication of hybrid photoelectrodes on a subsecond timescale with low energy consumption and possessing high photocurrent densities remains a centerpiece for successful implementation of photoelectrocatalytic synthesis of fuels and value-added chemicals. Here, we introduce a laser-driven technology to print sensitizers with desired morphologies and layer thickness onto different substrates, such as glass, carbon, or carbon nitride (CN). The specially designed process uses a thin polymer reactor impregnated with transition metal salts, confining the growth of transition metal oxide (TMO) nanostructures on the interface in milliseconds, while their morphology can be tuned by the laser. Multiple nano-p-n junctions at the interface increase the electron/hole lifetime by efficient charge trapping. A hybrid copper oxide/CN photoanode with optimal architecture reaches 10 times higher photocurrents than the pristine CN photoanode. This technology provides a modular approach to build a library of TMO-based composite films, enabling the creation of materials for diverse applications., Fabrication of hybrid photoelectrodes on a subsecond timescale with low energy consumption remains a challenge. Here, the authors report a modular approach, laser-driven transfer synthesis, to build a library of structurally defined transition metal oxide composite films, enabling the creation of materials for diverse applications.

The use of organic materials with reversible redox activity holds enormous potential for next‐generation Li‐ion energy storage devices. Yet, most candidates are not truly sustainable, i.e., not derived from renewable feedstock or made in benign reactions. Here an attempt is reported to resolve this issue by synthesizing an organic cathode material from tannic acid and microporous carbon derived from biomass. All constituents, including the redox‐active material and conductive carbon additive, are made from renewable resources. Using a simple, sustainable fabrication method, a hybrid material is formed. The low cost and ecofriendly material shows outstanding performance with a capacity of 108 mAh g−1 at 0.1 A g−1 and low capacity fading, retaining approximately 80% of the maximum capacity after 90 cycles. With approximately 3.4 V versus Li+/Li, the cells also feature one of the highest reversible redox potentials reported for biomolecular cathodes. Finally, the quinone‐catecholate redox mechanism responsible for the high capacity of tannic acid is confirmed by electrochemical characterization of a model compound similar to tannic acid but without catecholic groups., Tannic acid and sucrose‐derived porous carbon are combined to create a hybrid material using ball milling. The sustainable hybrid material shows exceptional properties as a cathode material for lithium‐ion energy storage devices that combines energy storage using both a redox process, from tannic acid, and electric double‐layer capacitance, from the porous carbon., DFG http://dx.doi.org/10.13039/100004807, German Federal Ministry of Education and Research, BMBF http://dx.doi.org/10.13039/501100002347, Fraunhofer‐Max Planck cooperation project, Max Planck Society http://dx.doi.org/10.13039/501100004189, Max Planck Institute of Colloids and Interfaces, Deutsche Forschungsgemeinschaft http://dx.doi.org/10.13039/501100001659

Short-lived micro-sized thermal gradients are challenging to measure. Especially, in thin film processes and devices, it is important to know the exact temperature profile to assure process parameters and the stability of sensitive materials. Many theoretical models try to describe the occurring temperatures, but still lack in profound experimental data. Here, a facile approach is presented, which allows to measure confined temperature gradients with millisecond and micrometer precision. By casting a thin alkane film onto a substrate of interest, it is possible to reconstruct local temperature gradients by imaging the phase behavior and morphology of the alkane film with a simple optical microscope setup. Alkanes are inert and their melting and boiling temperatures depend on the chain length. This allows to measure temperatures between 37 and 522 °C on any surface. Furthermore, after thorough characterization of laser-induced temperature gradients, this approach can be used to measure the phase transition behavior of complex thin film polymer mixtures.

SARS-CoV-2 is the causative agent of COVID-19 and is a severe threat to global health. Patients infected with SARS-CoV-2 show a wide range of symptoms and disease severity, while limited data is available on its immunogenicity.Here, the kinetics of the development of SARS-CoV-2-specific antibody responses in relation to clinical features and dynamics of specific B-cell populations are reported. Immunophenotyping of B cells was performed by flow cytometry with longitudinally collected PBMCs. In parallel, serum samples were analyzed for the presence of SARS-CoV-2-specific IgA, IgG, and IgM antibodies using whole proteome peptide microarrays. Soon after disease onset in a mild case, we observed an increased frequency of plasmablasts concomitantly with a strong SARS-CoV-2-specific IgA response. In contrast, a case with more severe progression showed a delayed, but eventually very strong and broad SARS-CoV-2-specific IgA response.This case study shows that determining SARS-CoV-2-specific antibody epitopes can be valuable to monitor the specificity and magnitude of the early B-cell response, which could guide the development of vaccine candidates. Follow-up studies are required to evaluate whether the kinetics and strength of the SARS-CoV-2-specific IgA response could be potential prognostic markers of viral control.

Single glycan–protein interactions are often weak, such that glycan binding partners commonly utilize multiple, spatially defined binding sites to enhance binding avidity and specificity. Current array technologies usually neglect defined multivalent display. Laser‐based array synthesis technology allows for flexible and rapid on‐surface synthesis of different peptides. By combining this technique with click chemistry, neo‐glycopeptides were produced directly on a functionalized glass slide in the microarray format. Density and spatial distribution of carbohydrates can be tuned, resulting in well‐defined glycan structures for multivalent display. The two lectins concanavalin A and langerin were probed with different glycans on multivalent scaffolds, revealing strong spacing‐, density‐, and ligand‐dependent binding. In addition, we could also measure the surface dissociation constant. This approach allows for a rapid generation, screening, and optimization of a multitude of multivalent scaffolds for glycan binding., Interactions of proteins with glycans rely on multivalency, where multiple adjacent binding events are involved. To study this glycan density‐dependent binding, a facile technique to synthesize peptide‐based multivalent carbohydrate scaffolds in parallel, directly on‐chip, in the microarray format, was developed. This allows to rapidly generate arrays of multivalent glycan structures, without the need for individual multistep syntheses per compound.

In this work, we present a qualitative and quantitative experimental analysis, as well as a numerical model, of a novel variant of the laser-induced forward transfer, which uses millisecond laser pulses. In this process, soft material nanolayer spots are transferred from a donor slide, which is coated with the soft material layer, to an acceptor slide via laser irradiation. This method offers a highly flexible material transfer to perform high-throughput combinatorial chemistry for the generation of biomolecule arrays. For the first time, we show visual evidence that the main transfer mechanism is contact-based, due to thermal surface expansion of the donor layer. Thus, the process is different from the many known variants of laser-induced forward transfer. We characterize the maximum axial surface expansion in relation to laser power and pulse duration. On this basis, we derive a numerical model that approximates the axial surface expansion within measurement tolerances. Finally, we analyze the topology of the transferred soft material nanolayer spots by fluorescence imaging and vertical scanning interferometry to determine width, height, and shape of the transferred material. Concluding from this experimental and numerical data, we can now predict the amount of transferred material in this process.

Emergence and re-emergence of pathogens bearing the risk of becoming a pandemic threat are on the rise. Increased travel and trade, growing population density, changes in urbanization, and climate have a critical impact on infectious disease spread. Currently, the world is confronted with the emergence of a novel coronavirus SARS-CoV-$_{2}$, responsible for yet more than 800 000 deaths globally. Outbreaks caused by viruses, such as SARS-CoV-$_{2}$, HIV, Ebola, influenza, and Zika, have increased over the past decade, underlining the need for a rapid development of diagnostics and vaccines. Hence, the rational identification of biomarkers for diagnostic measures on the one hand, and antigenic targets for vaccine development on the other, are of utmost importance. Peptide microarrays can display large numbers of putative target proteins translated into overlapping linear (and cyclic) peptides for a multiplexed, high-throughput antibody analysis. This enabled for example the identification of discriminant/diagnostic epitopes in Zika or influenza and mapping epitope evolution in natural infections versus vaccinations. In this review, we highlight synthesis platforms that facilitate fast and flexible generation of high-density peptide microarrays. We further outline the multifaceted applications of these peptide array platforms for the development of serological tests and vaccines to quickly encounter pandemic threats.

Poly(dimethylacrylamide) (PDMA) based matrix materials were developed for laser-based in situ solid phase peptide synthesis to produce high density arrays. In this specific array synthesis approach, amino acid derivatives are embedded into a matrix material, serving as a “solid” solvent material at room temperature. Then, a laser pulse transfers this mixture to the target position on a synthesis slide, where the peptide array is synthesized. Upon heating above the glass transition temperature of the matrix material, it softens, allowing diffusion of the amino acid derivatives to the synthesis surface and serving as a solvent for peptide bond formation. Here, we synthesized PDMA six-arm star polymers, offering the desired matrix material properties, using atom transfer radical polymerization. With the synthesized polymers as matrix material, we structured and synthesized arrays with combinatorial laser transfer. With densities of up to 20,000 peptide spots per cm 2 , the resolution could be increased compared to the commercially available standard matrix material. Time-of-Flight Secondary Ion Mass Spectrometry experiments revealed the penetration behavior of an amino acid derivative into the prepared acceptor synthesis surface and the effectiveness of the washing protocols.

We developed a next-generation method for chemical in–situ combinatorial biomolecule array synthesis. This allows for an unprecedented combinatorial freedom in the automated chemical synthesis of molecule arrays with very high spot densities. Key feature of this new method is an automated positioning and laser transfer process: Small solid material spots are rapidly transferred from a donor film to an acceptor surface, requiring only minute amounts of materials. The transfer is performed with different and easy-to-produce donor slides. Each donor slide bears a thin polymer film, embedding one type of monomer. The coupling reaction occurs in a separate heating step, where the matrix becomes viscous and building blocks can diffuse within the material and couple to the acceptor surface. Since these transferred material spots are only several nanometers thin, this method allows for a consecutive multi-layer material deposition of e.g. activation reagents and amino acids. Subsequent heat-induced mixing facilitates an in–situ activation and coupling of the monomers. Positioning several of such resin spots, containing different chemical reagents, on top of each other, will enable for the first time in such small dimensions unique chemical synthesis strategies for each spot. Amount and concentration of the deposited materials can be adjusted with the laser parameters. Employing similar arrays, we can analyze the human immune response towards the proteome of different pathogens. We screened several peptide array replicas with different patient sera. The screenings resulted in significant hits in several proteins with interesting implications for future diagnostics and vaccine development.

The Ebola virus (EBOV) can cause severe infections in humans, leading to a fatal outcome in a high percentage of cases. Neutralizing antibodies against the EBOV surface glycoprotein (GP) can prevent infections, demonstrating a straightforward way for an efficient vaccination strategy. Meanwhile, many different anti-EBOV antibodies have been identified, whereas the exact binding epitopes are often unknown. Here, we present the analysis of serum samples from an EBOV vaccine trial with the viral vector vaccine rVSV-ZEBOV and an Ebola virus disease (EVD) survivor, using high-density peptide arrays. In this proof-of-principle study, we detected distinct IgG and IgM antibodies binding to different epitopes of EBOV GP: By mapping the whole GP as overlapping peptide fragments, we found new epitopes and confirmed epitopes from the literature. Furthermore, we could validate the highly selective binding epitope of a neutralizing monoclonal anti-EBOV GP antibody. This shows that peptide arrays can be a valuable tool to study the humoral immune response to vaccines in patients and to support Ebola vaccine development. This article is protected by copyright. All rights reserved

Glycan microarrays have become a powerful technology to study biological processes, such as cell-cell interaction, inflammation, and infections. Yet, several challenges, especially in multivalent display, remain. In this introductory lecture we discuss the state-of-the-art glycan microarray technology, with emphasis on novel approaches to access collections of pure glycans and their immobilization on surfaces. Future directions to mimic the natural glycan presentation on an array format, as well as in situ generation of combinatorial glycan collections, are discussed.

Here, the combinatorial synthesis of molecule arrays via a laser-assisted process is reported. Laser-transferred polymer nanolayers with embedded monomers, activators, or bases can be reliably stacked on top of each other, spot-by-spot, to synthesize molecule arrays. These various chemicals in the nanometer-thin layers are mixed by heat or solvent vapor, inducing coupling reactions. As an example, peptoid arrays with a density of 10 000 spots per cm2 with the sub-monomer or monomer method are generated. Moreover, successful reactions spot-by-spot are verified by laser-transferring MALDI-matrix (Matrix-assisted laser desorption/ionization) followed by MALDI mass spectrometry imaging.

In this review, we describe different methods of microarray fabrication based on the use of micro-particles/-beads and point out future tendencies in the development of particle-based arrays. First, we consider oligonucleotide bead arrays, where each bead is a carrier of one specific sequence of oligonucleotides. This bead-based array approach, appearing in the late 1990s, enabled high-throughput oligonucleotide analysis and had a large impact on genome research. Furthermore, we consider particle-based peptide array fabrication using combinatorial chemistry. In this approach, particles can directly participate in both the synthesis and the transfer of synthesized combinatorial molecules to a substrate. Subsequently, we describe in more detail the synthesis of peptide arrays with amino acid polymer particles, which imbed the amino acids inside their polymer matrix. By heating these particles, the polymer matrix is transformed into a highly viscous gel, and thereby, imbedded monomers are allowed to participate in the coupling reaction. Finally, we focus on combinatorial laser fusing of particles for the synthesis of high-density peptide arrays. This method combines the advantages of particles and combinatorial lithographic approaches.

A low-cost laser-based printing setup is presented, which allows for the spot-wise patterning of surfaces with defined polymer nanolayers. These nanolayer spots serve as a “solid solvent,” embedding different chemicals, chemical building blocks, materials, or precursors and can be stacked on top of each other. By melting the spot pattern, the polymer-embedded molecules are released for chemical reaction. This enables researchers to quickly pattern a surface with different molecules and materials, mixing them directly on the surface for high-throughput chemical synthesis to generate and screen diverse microarray libraries. In contrast to expensive ink-jet or contact printing, this approach does not require premixing of inks, which enables in situ combinatorial mixing. Easy access and versatility of this patterning approach are shown by generating microarrays of various biomolecules, such as glycans for the first time, to screen interactions of antibodies and lectins. In addition, a layer-by-layer solid-phase synthesis of peptides directly on the microarray is presented. Amino acid–containing nanolayers are repeatedly laser-transferred and reacted with the functionalized acceptor surface in defined patterns. This simple system enables a reproducible array production, down to spot-to-spot distances of 100 μm, and offers a flexible and cheap alternative to expensive spotting robot technology.

Lyme disease is the most common tick-borne infectious disease in Europe and North America. Previous studies discovered the immunogenic role of a surface-exposed lipoprotein (VlsE) of Borreliella burgdorferi. We employed high density peptide arrays to investigate the antibody response to the VlsE protein in VlsE-positive patients by mapping the protein as overlapping peptides and subsequent in-depth epitope substitution analyses. These investigations led to the identification of antibody fingerprints represented by a number of key residues that are indispensable for the binding of the respective antibody. This approach allows us to compare the antibody specificities of different patients to the resolution of single amino acids. Our study revealed that the sera of VlsE-positive patients recognize different epitopes on the protein. Remarkably, in those cases where the same epitope is targeted, the antibody fingerprint is almost identical. Furthermore, we could correlate two fingerprints with human autoantigens and an Epstein-Barr virus epitope; yet, the link to autoimmune disorders seems unlikely and must be investigated in further studies. The other three fingerprints are much more specific for B. burgdorferi. Since antibody fingerprints of longer sequences have proven to be highly disease specific, our findings suggest that the fingerprints could function as diagnostic markers that can reduce false positive test results. This article is protected by copyright. All rights reserved

B-cell epitope sequences from Zika virus (ZIKV) NS1 protein have been identified using epitope prediction tools. Mapping these sequences onto the NS1 surface reveals two major conformational epitopes and a single linear one. Despite an overall average sequence identity of ca. 55% between the NS1 from ZIKV and the four dengue virus (DENV) serotypes, epitope sequences were found to be highly conserved. Nevertheless, nonconserved epitope-flanking residues are responsible for a dramatically divergent electrostatic surface potential on the epitope regions of ZIKV and DENV2 serotypes. These findings suggest that strategies for differential diagnostics on the basis of short linear NS1 sequences are likely to fail due to immunological cross-reactions. Overall, results provide the molecular basis of differential discrimination between Zika and DENVs by NS1 monoclonal antibodies.

The antibody species that patrol in a patient's blood are an invaluable part of the immune system. While most of them shield us from life-threatening infections, some of them do harm in autoimmune diseases. If we knew exactly all the antigens that elicited all the antibody species within a group of patients, we could learn which ones correlate with immune protection, are irrelevant, or do harm. Here, we demonstrate an approach to this question: First, we use a plethora of phage-displayed peptides to identify many different serum antibody binding peptides. Next, we synthesize identified peptides in the array format and rescreen the serum used for phage panning to validate antibody binding peptides. Finally, we systematically vary the sequence of validated antibody binding peptides to identify those amino acids within the peptides that are crucial for binding "their" antibody species. The resulting immune fingerprints can then be used to trace them back to potential antigens. We investigated the serum of an individual in this pipeline, which led to the identification of 73 antibody fingerprints. Some fingerprints could be traced back to their most likely antigen, for example the immunodominant capsid protein VP1 of enteroviruses, most likely elicited by the ubiquitous poliovirus vaccination. Thus, with our approach, it is possible, to pinpoint those antibody species that correlate with a certain antigen, without any pre-information. This can help to unravel hitherto enigmatic diseases.

The development of an efficacious and practicable vaccine conferring sterile immunity towards a Plasmodium infection represents a not yet achieved goal. A crucial factor for the impact of a given anti-plasmodial subunit vaccine is the identification of the most potent parasitic components required to induce protection from both infection and disease. Here, we present a method based on a novel high-density peptide array technology that allows for a flexible readout of malaria antibodies. Peptide arrays applied as a screening method can be used to identify novel immunogenic antibody epitopes under a large number of potential antigens/peptides. Ultimately, discovered antigen candidates and/or epitope sequences can be translated into vaccine prototype design. The technology can be further utilized to unravel antibody-mediated immune responses (e.g., involved in the establishment of semi-immunity) and moreover to confirm vaccine potency during the process of clinical development by verifying the induced antibody responses following vaccination.

Biofunctionalization of surfaces in a microarray format has revolutionized biological assay applications. Here, a microarray system based on a microelectronic chip is presented that allows for a versatile combinatorial in situ molecule synthesis with very high density. Successfully demonstrating an application for peptide array synthesis, the method offers a compact approach, high combinatorial freedom, and, due to the intrinsic alignment, high and reproducible precision. Patterning the chip surface with different microparticle types which imbed different monomers, several thousand different molecule types can be simultaneously elongated layer-by-layer by coupling the particle imbedded monomers to the molecules growing on the chip surface. This technique has the potential for a wide application in combinatorial chemistry, as long as the desired monomeric building blocks are compatible with the chemical process.

Surface-bound microarrays of multiple oligo- and macromolecules (e.g., peptides, DNA) offer versatile options in biomedical applications like drug screening, DNA analysis, or medical diagnostics. Combinatorial syntheses of these molecules in situ can save significant resources in regard to processing time and material use. Furthermore, high feature densities are needed to enable high-throughput and low sample volumes as generally regarded in combinatorial chemistry. Here, a scanning-probe-lithography-based approach for the combinatorial in situ synthesis of macromolecules is presented in microarray format. Feature sizes below 40 µm allow for the creation of high-density arrays with feature densities of 62 500 features per cm2 . To demonstrate feasibility of this approach for biomedical applications, a multiplexed array of functional protein tags (HA- and FLAG-tag) is synthesized, and selective binding of respective epitope recognizing antibodies is shown. This approach uses only small amounts of base chemicals for synthesis and can be further parallelized, therefore, opening up a route to flexible, highly dense, and cost-effective microarrays.