Controlling Nanoparticle Distance by On-Surface DNA-Origami Folding.

DNA origami is a flexible platform for the precise organization of nano-objects, enabling numerous applications from biomedicine to nano-photonics. Its huge potential stems from its high flexibility that allows customized structures to meet specific requirements. The ability to generate diverse final structures from a common base by folding significantly enhances design variety and is regularly occurring in liquid. This study describes a novel approach that combines top-down lithography with bottom-up DNA origami techniques to control folding of the DNA origami with the adsorption on pre-patterned surfaces. Using this approach, tunable plasmonic dimer nano-arrays are fabricated on a silicon surface. This involves employing electron beam lithography to create adsorption sites on the surface and utilizing self-organized adsorption of DNA origami functionalized with two gold nanoparticles (AuNPs). The desired folding of the DNA origami helices can be controlled by the size and shape of the adsorption sites. This approach can for example be used to tune the center-to-center distance of the AuNPs dimers on the origami template. To demonstrate this technique's efficiency, the Raman signal of dye molecules (carboxy tetramethylrhodamine, TAMRA) coated on the AuNPs surface are investigated. These findings highlight the potential of tunable DNA origami-based plasmonic nanostructures for many applications.

Positional control of DNA origami based gold dimer hybrid nanostructures on pre-structured surfaces.

This study explores important parameters for achieving a high-level positional control of DNA-nanoparticle hybrid structures by drop-casting onto a pre-structured silicon surface, in which the active adsorption sites were defined using electron beam lithography. By confining the adsorption sites to the scale of the DNA origami, we create multi-dimensional patterns and study the effect of diffusion and hybrid nanostructure concentration in the liquid on site occupation. We also propose a physical diffusion model that highlights the importance of surface diffusion in facilitating the adsorption of hybrid nanostructure onto active sites, particularly for two and one-dimensional adsorption sites. Our study shows prominent results of the hybrid nanostructure's selective adsorption, indicating high adsorption efficiency and precise control over the position, as well as the spatial orientation. We anticipate similar results in related systems, both in terms of different surfaces and similar DNA structures. Overall, our findings offer promising prospects for the development of large-scale nanoarrays on micrometer-scale surfaces with nanometer precision and orientation control.

Biomacromolecules as tools and objects in nanometrology-current challenges and perspectives.

Nucleic acids, proteins, and polysaccharides are the most important classes of biopolymers. The inherent properties of biomacromolecules are contrary to those of well-defined small molecules consequently raising a number of specific challenges which become particularly apparent if biomacromolecules are treated as objects in quantitative analysis. At the same time, their specific functional ability of molecular recognition and self-organization (e.g., enzymes, antibodies, DNA) enables us to make biomacromolecules serving as molecular tools in biochemistry and molecular biology, or as precisely controllable dimensional platforms for nanometrological applications. Given the complexity of biomacromolecules, quantitative analysis is not limited to the measurement of their concentration but also involves the determination of numerous descriptors related to structure, interaction, activity, and function. Among the biomacromolecules, glycans set examples that quantitative characterization is not necessarily directed to the measurement of amount-of-substance concentration but instead involves the determination of relative proportions (molar ratios) of structural features for comparison with theoretical models. This article addresses current activities to combine optical techniques such as Raman spectroscopy with isotope dilution approaches to realize reference measurement procedures for the quantification of protein biomarkers as an alternative to mass spectrometry-based techniques. Furthermore, it is explored how established ID-MS protocols are being modified to make them applicable for quantifying virus proteins to measure the HIV viral load in blood samples. As an example from the class of carbohydrates, the challenges in accurate determination of substitution patterns are outlined and discussed. Finally, it is presented that biomacromolecules can also serve as tools in quantitative measurements of dimensions with an example of DNA origami to generate defined dimensional standards to be used for calibration in super-resolution fluorescence microscopy. Graphical abstract Quantitative analysis of biomacromolecules is accompanied with special challenges different from low molecular weight compounds. In addition, they are not only objects but also tools applicable for quantitative measurements.

Comparison of potential higher order reference methods for total haemoglobin quantification-an interlaboratory study.

The total haemoglobin (Hb) concentration in blood is one of the most frequently measured analytes in clinical medicine because of its significance for evaluating the health state of a human. The spectrophotometric cyanmethaemoglobin (HiCN) method is the internationally accepted conventional reference method to determine this biomarker. It is frequently used in clinical routine diagnostics but is not traceable to the International System of Units and thus does not meet highest metrological demands. A further critical issue is the toxicity of the necessary potassium cyanide. Different methods to solve these problems are reported here. They all were validated against the HiCN method in an interlaboratory comparison by measuring the total Hb concentration present in the certified reference material JCCRM 912-2M. Methods considered were the spectrophotometric alkaline haematin detergent (AHD) method as well as several isotope dilution (ID)-based approaches. The latter include inductively coupled plasma mass spectrometry (ICP-MS), species-specific (SS) ICP-MS, organic MS and Raman spectrometry. Graphical abstract ᅟ.

Direct Growth of Patterned Graphene.

The direct growth of single-layer graphene patterns via electron irradiation of aromatic self-assembled monolayers and subsequent annealing is demonstrated. In this way, a reduction in the number of necessary manufacturing steps is achieved. The formed micro- and nanostructures can be arbitrarily shaped and eventually implemented in a manifold of applications.

State of the art Raman techniques for biological applications.

Raman spectroscopy is a powerful tool for the elucidation of qualitative and quantitative information from biological systems and has huge potential in areas such as biotechnologies, drug discovery, agro-chemical research and clinical diagnostics. This report summarises the principal Raman techniques applied to biomedical systems and discusses the challenges that exist to the wide spread adoption of Raman spectroscopy.

Optical Tomography of Polymeric Microsphere Layers Using Confocal Raman Microscopy.

In confocal Raman microscopy, depth profiling is a key application that enables analysis of the structural and chemical composition and size of three-dimensional (3D) transparent objects. However, the precise interpretation of a probed sample's Raman depth profile measurement can be significantly affected by both its size and surrounding objects. This study provides a more comprehensive understanding of the observed optical effects at the interface between polymer spheres and different substrates. Ray- and wave-optical simulations support our results. We derive a correction factor that, depending on the instrumental configuration, allows us to determine the nominal dimensions of the scanned objects more accurately from Raman depth profiles. Our studies support the need for careful consideration when employing depth profiling in confocal Raman microscopy for nondestructive, quantitative tomography of 3D objects.

Confocal Raman Microscopy for the Analysis of the Three-Dimensional Shape of Polymeric Microsphere Layers.

The reconstruction of the three-dimensional (3D) morphology of polymeric microsphere layers based on confocal Raman microscopy was studied. Refraction of the Raman laser beam at the curved surface of the spheres broadens the focus volume inside the sphere. Compared to planar layers, the focus gets trapped inside the spheres such that the measured depth profiles are shifted and broadened. Additionally, the Raman signal of the underlying substrate is already observed for nominal focus positions above the microsphere layer. The results are successfully modeled with ray-optical simulations that allow for a clear understanding of the relevant mechanisms that lead to the generation of the Raman signals in the complex three-dimensional structures.

Impact of Polymer-Assisted Epitaxial Graphene Growth on Various Types of SiC Substrates.

The growth parameters for epitaxial growth of graphene on silicon carbide (SiC) have been the focus of research over the past few years. However, besides the standard growth parameters, the influence of the substrate pretreatment and properties of the underlying SiC wafer are critical parameters for optimizing the quality of monolayer graphene on SiC. In this systematic study, we show how the surface properties and the pretreatment determine the quality of monolayer graphene using polymer-assisted sublimation growth (PASG) on SiC. Using the spin-on deposition technique of PASG, several polymer concentrations have been investigated to understand the influence of the polymer content on the final monolayer coverage using wafers of different miscut angles and different polytypes. Confocal laser scanning microscopy (CLSM), atomic force microscopy (AFM), Raman spectroscopy, and scanning electron microscopy (SEM) were used to characterize these films. The results show that, even for SiC substrates with high miscut angles, high-quality graphene is obtained when an appropriate polymer concentration is applied. This is in excellent agreement with the model understanding that an insufficient carbon supply from SiC step edge decomposition can be compensated by additionally providing carbon from a polymer source. The described methods make the PASG spin-on deposition technique more convenient for commercial use.

Mechanically stacked 1-nm-thick carbon nanosheets: ultrathin layered materials with tunable optical, chemical, and electrical properties.

Carbon nanosheets are mechanically stable, free-standing two-dimensional materials with a thickness of ≈1 nm and well defined physical and chemical properties. They are made by radiation-induced cross-linking of aromatic self-assembled monolayers. Herein, a route is presented to the scalable fabrication of multilayer nanosheets with tunable electrical, optical, and chemical properties on insulating substrates. Stacks of up to five nanosheets with sizes of ≈1 cm(2) on oxidized silicon are studied. Their optical characteristics are investigated by visual inspection, optical microscopy, UV-vis reflection spectroscopy, and model calculations. Their chemical composition is studied by X-ray photoelectron spectroscopy. The multilayer samples are then annealed in an ultrahigh vacuum at various temperatures up to 1100 K. A subsequent investigation by Raman, X-ray photoelectron, and UV-vis reflection spectroscopy, as well as by electrical four-point probe measurements, demonstrates that the layered nanosheets transform into nanocrystalline graphene. This structural and chemical transformation is accompanied by changes in the optical properties and electrical conductivity and opens up a new path for the fabrication of ultrathin functional conductive coatings.

Double isotope dilution surface-enhanced Raman scattering as a reference procedure for the quantification of biomarkers in human serum.

Double isotope dilution surface-enhanced Raman scattering (double IDSERS) is qualified as a method for accurate and precise determination of biomarkers in human blood serum. Providing a full evaluation of the measurement uncertainty as well as traceability to a reference material sets the procedure in line with the requirements of a primary ratio method. Data evaluation is based on a partial least squares (PLS) model, whose prediction ability is validated from quantifying the uric acid concentration in both an artificial reference solution and a real human blood serum sample. With the proposed approach, the uric acid serum concentration can be determined with an uncertainty of 1.6% at a confidence level of 95%.

Step-by-step monitoring of a magnetic and SERS-active immunosensor assembly for purification and detection of tau protein.

We report a bottom-up synthesis of iron oxide and gold nanoparticles, which are functionalized and combined to form a nanohybrid serving as an immune sensor, which selectively binds to tau protein, a biomarker for diagnosis of Alzheimer's disease. Detection of the target analyte is achieved by surface-enhanced Raman scattering originating from the diagnostic part of the nanohybrid that was prepared from Au nanoparticles functionalized with 5,5'-dithiobis-(2-nitrobenzoic acid) as a Raman reporter and monoclonal anti-tau antibody. The magnetic part consists of Fex Oy nanoparticles functionalized with polyclonal anti-tau antibody and is capable to separate tau protein from a complex matrix such as cerebrospinal fluid. We further identified and validated a set of analytical tools that allow monitoring the success of both nanoparticle preparation and each functionalization step performed during the assembly of the two binding sites by an immune reaction. By applying UV/Vis spectroscopy, dynamic light scattering, zeta potential measurements, X-ray diffraction, small-angle X-ray scattering, and transmission electron microscopy, we demonstrate a proof-of-concept for a controlled and step-by-step traceable synthesis of a tau protein-specific immune sensor.

CO2 laser-induced zonation in dental enamel: a Raman and IR microspectroscopic study.

The gradient of structural alteration and molecular exchange across CO(2) laser-irradiated areas in dental enamel was analyzed by Raman and attenuated total reflectance infrared microspectroscopy. The type and the degree of structural changes in morphologically distinguishable zones within the laser spot vary depending on the laser-irradiation parameters--power (1 and 3 W), treatment time (5 and 10 s), and operational mode (super pulse and continuous wave). Using higher power, irrespective of the operation mode, the enamel tissue ablates and a crater is formed. The prevalent phase at the bottom of the crater is dehydrated O(2) (2-)-bearing apatite, that is, the fundamental framework topology is preserved. Additional nonapatite calcium phosphate phases are located mainly at the slope of the laser crater. No structural transformation of mineral component was detected aside the crater rim, only a CO(3)-CO(2) exchange, which decays with the radial distance. A lower-power laser irradiation slightly roughens the enamel surface and the structural modification of enamel apatite is considerably weaker for continuous wave than for super pulse mode. Prolonged low-power laser treatment results in recrystallization, and thus structural recovering of apatite might be of clinical relevance for enamel surface treatments.

Traceable Quantitative Raman Microscopy and X-ray Fluorescence Analysis as Nondestructive Methods for the Characterization of Cu(In,Ga)Se2 Absorber Films.

The traceability of measured quantities is an essential condition when linking process control parameters to guaranteed physical properties of a product. Using Raman spectroscopy as an analytical tool for monitoring the production of Cu(In1-xGax)Se2 thin-film solar cells, proper calibration with regard to chemical composition and lateral dimensions is a key prerequisite. This study shows how the multiple requirements of calibration in Raman microscopy might be addressed. The surface elemental composition as well as the integral elemental composition of the samples is traced back by reference-free X-ray fluorescence analysis. Reference Raman spectra are then generated for the relevant Cu(In1-xGax)Se2 related compounds. The lateral dimensions are calibrated with the help of a novel dimensional standard whose regular structures have been traced back to the International System of Units by metrological scanning force microscopy. On this basis, an approach for the quantitative determination of surface coverage values from lateral Raman mappings is developed together with a complete uncertainty budget. Raman and X-ray spectrometry have here been proven as complementary nondestructive methods combining surface sensitivity and in-depth information on elemental and species distribution for the reliable quality control of Cu(In1-xGax)Se2 absorbers and Cu(In1-xGax)3Se5 surface layer formation.

Epitaxial graphene on SiC: modification of structural and electron transport properties by substrate pretreatment.

The electrical transport properties of epitaxial graphene layers are correlated with the SiC surface morphology. In this study we show by atomic force microscopy and Raman measurements that the surface morphology and the structure of the epitaxial graphene layers change significantly when different pretreatment procedures are applied to nearly on-axis 6H-SiC(0 0 0 1) substrates. It turns out that the often used hydrogen etching of the substrate is responsible for undesirable high macro-steps evolving during graphene growth. A more advantageous type of sub-nanometer stepped graphene layers is obtained with a new method: a high-temperature conditioning of the SiC surface in argon atmosphere. The results can be explained by the observed graphene buffer layer domains after the conditioning process which suppress giant step bunching and graphene step flow growth. The superior electronic quality is demonstrated by a less extrinsic resistance anisotropy obtained in nano-probe transport experiments and by the excellent quantization of the Hall resistance in low-temperature magneto-transport measurements. The quantum Hall resistance agrees with the nominal value (half of the von Klitzing constant) within a standard deviation of 4.5 × 10(-9) which qualifies this method for the fabrication of electrical quantum standards.

All-carbon vertical van der Waals heterostructures: non-destructive functionalization of graphene for electronic applications.

Non-destructive chemical functionalization of graphene for applications in electronic devices (e.g., sensors or transducers) is achieved via assembly of carbon nanomembrane (CNM)/single-layer graphene (SLG) van der Waals heterostructures. The CNMs are 1 nm-thick, dielectric molecular sheets terminated with functional amino groups. The structure and performance of heterostructured field-effect transistors (FETs) are characterized by photoelectron/Raman spectroscopy and by electric transport measurements in vacuum, ambient conditions and water.

Functional single-layer graphene sheets from aromatic monolayers.

Self-assembled monolayers of aromatic molecules on copper substrates can be converted into high-quality single-layer graphene using low-energy electron irradiation and subsequent annealing. This two-dimensional solid state transformation is characterized on the atomic scale and the physical and chemical properties of the formed graphene sheets are studied by complementary microscopic and spectroscopic techniques and by electrical transport measurements. As substrates, Cu(111) single crystals and the technologically relevant polycrystalline copper foils are successfully used.

On-chip approach for traceable quantification of biomarkers based on isotope-dilution surface-enhanced Raman scattering (IDSERS).

We present an on-chip approach for the quantification of biomarkers based on isotope-dilution surface-enhanced Raman scattering (IDSERS). The full procedure was realized on a few square millimetres of a SERS-active substrate, covered with either lithographically engineered gold nanotriangles or silver nanospheres generated by galvanic displacement deposition. The use of certified reference materials combined with the ID principle ensures traceability of the quantitation to SI units. A series of substance spots was deposited onto the SERS active area and measured one by one in fast sequence. The SERS spectra were used to generate and validate a PLS model and also to predict the creatinine concentration of an unknown serum sample.

Conversion of self-assembled monolayers into nanocrystalline graphene: structure and electric transport.

Graphene-based materials have been suggested for applications ranging from nanoelectronics to nanobiotechnology. However, the realization of graphene-based technologies will require large quantities of free-standing two-dimensional (2D) carbon materials with tunable physical and chemical properties. Bottom-up approaches via molecular self-assembly have great potential to fulfill this demand. Here, we report on the fabrication and characterization of graphene made by electron-radiation induced cross-linking of aromatic self-assembled monolayers (SAMs) and their subsequent annealing. In this process, the SAM is converted into a nanocrystalline graphene sheet with well-defined thickness and arbitrary dimensions. Electric transport data demonstrate that this transformation is accompanied by an insulator to metal transition that can be utilized to control electrical properties such as conductivity, electron mobility, and ambipolar electric field effect of the fabricated graphene sheets. The suggested route opens broad prospects toward the engineering of free-standing 2D carbon materials with tunable properties on various solid substrates and on holey substrates as suspended membranes.