In Vitro and In Silico Studies of Functionalized Polyurethane Surfaces toward Understanding Biologically Relevant Interactions.

The solid-aqueous boundary formed upon biomaterial implantation provides a playground for most biochemical reactions and physiological processes involved in implant-host interactions. Therefore, for biomaterial development, optimization, and application, it is essential to understand the biomaterial-water interface in depth. In this study, oxygen plasma-functionalized polyurethane surfaces that can be successfully utilized in contact with the tissue of the respiratory system were prepared and investigated. Through experiments, the influence of plasma treatment on the physicochemical properties of polyurethane was investigated by atomic force microscopy, attenuated total reflection infrared spectroscopy, differential thermal analysis, X-ray photoelectron spectroscopy, secondary ion mass spectrometry, and contact angle measurements, supplemented with biological tests using the A549 cell line and two bacteria strains (Staphylococcus aureus and Pseudomonas aeruginosa). The molecular interpretation of the experimental findings was achieved by molecular dynamics simulations employing newly developed, fully atomistic models of unmodified and plasma-functionalized polyurethane materials to characterize the polyurethane-water interfaces at the nanoscale in detail. The experimentally obtained polar and dispersive surface free energies were consistent with the calculated free energies, verifying the adequacy of the developed models. A 20% substitution of the polymeric chain termini by their oxidized variants was observed in the experimentally obtained plasma-modified polyurethane surface, indicating the surface saturation with oxygen-containing functional groups.

Surface engineering of mixed conjugated/polyelectrolyte brushes - Tailoring interface structure and electrical properties.

HYPOTHESIS: Mixed polymer brushes (MPBs) could be synthesized by surface dilution of homopolymer brushes and subsequent grafting of other type of chains in the formed voids. Nanophase separation and dynamics of surface-grafted chains could be tailored by modification of their molecular architecture. Mixed polyelectrolyte and conjugated chains contribute synergistically to tailor properties of the coating. EXPERIMENTS: A new synthetic strategy that allowed spatially controlled grafting of poly(sodium 4-styrenesulfonate) chains (PSSNa) in close neighborhood of poly(3-methylthienyl methacrylate) (PMTM) brushes (precursors of the conjugated chains) using surface-initiated polymerizations was developed. The final mixed conjugated/polyelectrolyte brushes were prepared by template polymerization of pendant thiophene groups in PMTM chains. Surface dynamics and nanophase separation of MPBs were studied by nanoscale resolution IR imaging, SIMS profiling and AFM mapping in selective solvents. FINDINGS: Unconjugated MPBs were shown to undergo vertical, and horizontal nanophase separation, while the size and shape of the nanodomains were dependent on molar ratio of the mixed chains and their relative lengths. Generation of the conjugated chains led to diminishing of nanophase separation thanks to stronger mutual interactions of conjugated PMTM and PSSNa (macromolecular mixing). The obtained systems demonstrated tunable interfacial structure and resistance switching phenomenon desired in construction of smart surfaces or memristive devices.

Dendrite Formation on the Poly(methyl methacrylate) Surface Reactively Sputtered with a Cesium Ion Beam.

The development of topography plays an important role when low-energy projectiles are used to modify the surface or analyze the properties of various materials. It can be a feature that allows one to create complex structures on the sputtered surface. It can also be a factor that limits depth resolution in ion-based depth profiling methods. In this work, we have studied the evolution of microdendrites on poly(methyl methacrylate) sputtered with a Cs 1 keV ion beam. Detailed analysis of the topography of the sputtered surface shows a sea of pillars with islands of densely packed pillars, which eventually evolve to fully formed dendrites. The development of the dendrites depends on the Cs fluence and temperature. Analysis of the sputtered surface by physicochemical methods shows that the mechanism responsible for the formation of the observed microstructures is reactive ion sputtering. It originates from the chemical reaction between the target material and primary projectile and is combined with mass transport induced by ion sputtering. The importance of chemical reaction for the formation of the described structures is shown directly by comparing the change in the surface morphology under the same dose of a nonreactive 1 keV xenon ion beam.

Cell-Specific Response of NSIP- and IPF-Derived Fibroblasts to the Modification of the Elasticity, Biological Properties, and 3D Architecture of the Substrate.

The fibrotic fibroblasts derived from idiopathic pulmonary fibrosis (IPF) and nonspecific interstitial pneumonia (NSIP) are surrounded by specific environments, characterized by increased stiffness, aberrant extracellular matrix (ECM) composition, and altered lung architecture. The presented research was aimed at investigating the effect of biological, physical, and topographical modification of the substrate on the properties of IPF- and NSIP-derived fibroblasts, and searching for the parameters enabling their identification. Soft and stiff polydimethylsiloxane (PDMS) was chosen for the basic substrates, the properties of which were subsequently tuned. To obtain the biological modification of the substrates, they were covered with ECM proteins, laminin, fibronectin, and collagen. The substrates that mimicked the 3D structure of the lungs were prepared using two approaches, resulting in porous structures that resemble natural lung architecture and honeycomb patterns, typical of IPF tissue. The growth of cells on soft and stiff PDMS covered with proteins, traced using fluorescence microscopy, confirmed an altered behavior of healthy and IPF- and NSIP-derived fibroblasts in response to the modified substrate properties, enabling their identification. In turn, differences in the mechanical properties of healthy and fibrotic fibroblasts, determined using atomic force microscopy working in force spectroscopy mode, as well as their growth on 3D-patterned substrates were not sufficient to discriminate between cell lines.

Electrically Switchable Film Structure of Conjugated Polymer Composites.

Domains rich in different blend components phase-separate during deposition, creating a film morphology that determines the performance of active layers in organic electronics. However, morphological control either relies on additional fabrication steps or is limited to a small region where an external interaction is applied. Here, we show that different semiconductor-insulator polymer composites can be rapidly dip-coated with the film structure electrically switched between distinct morphologies during deposition guided by the meniscus formed between the stationary barrier and horizontally drawn solid substrate. Reversible and repeatable changes between the morphologies used in devices, e.g., lateral morphologies and stratified layers of semiconductors and insulators, or between phase-inverted droplet-like structures are manifested only for one polarity of the voltage applied across the meniscus as a rectangular pulse. This phenomenon points to a novel mechanism, related to voltage-induced doping and the doping-dependent solubility of the conjugated polymer, equivalent to an increased semiconductor content that controls the composite morphologies. This is effective only for the positively polarized substrate rather than the barrier, as the former entrains the nearby lower part of the coating solution that forms the final composite film. The mechanism, applied to the pristine semiconductor solution, results in an increased semiconductor deposition and 40-times higher film conductance.

Efficiency Boost in Dye-Sensitized Solar Cells by Post- Annealing UV-Ozone Treatment of TiO2 Mesoporous Layer.

The organic residues on titanium(IV) oxide may be a significant factor that decreases the efficiency of dye-sensitized solar cells (DSSC). Here, we suggest the UV-ozone cleaning process to remove impurities from the surface of TiO2 nanoparticles before dye-sensitizing. Data obtained from scanning electron microscopy, Kelvin probe, Fourier-transform infrared spectroscopy, and Raman spectroscopy showed that the amounts of organic contamination were successfully reduced. Additionally, the UV-VIS spectrophotometry, spectrofluorometry, and secondary ion mass spectrometry proved that after ozonization, the dyeing process was relevantly enhanced. Due to the removal of organics, the power conversion efficiency (PCE) of the prepared DSSC devices was boosted from 4.59% to 5.89%, which was mostly caused by the increment of short circuit current (Jsc) and slight improvement of the open circuit voltage (Voc).

Free-standing TiO2 nanotubes decorated with spherical nickel nanoparticles as a cost-efficient electrocatalyst for oxygen evolution reaction.

Here, we report significant activity towards the oxygen evolution reaction (OER) of spherical nickel nanoparticles (NPs) electrodeposited onto free-standing TiO2 nanotubes (TNT) via cyclic voltammetry. It has been shown that simple manipulation of processing parameters, including scan rate and number of cycles, allows for formation of the NPs in various diameters and amounts. The polarization data with respect to transmission electron microscopy (TEM) allowed for determination of the diameter and propagation depth of the Ni NPs leading to the highest activity towards the OER with an overpotential of 540 mV at +10 mA cm-2 and Tafel slope of 52 mV per decade. X-ray photoelectron spectroscopy (XPS) indicates the presence of structure defects within Ni NPs whereas Mott-Schottky analysis provides information on the anodically shifted flat band potential and highly increased donor density. The obtained results along with literature studies allowed a proposal of the origin of the enhancement towards the OER. We believe that combination of transition metal-based NPs and TNT provides valuable insight on efficient and low-cost electrocatalysts.

Biophysical and Biochemical Characteristics as Complementary Indicators of Melanoma Progression.

The multistep character of cancer progression makes it difficult to define a unique biomarker of the disease. Interdisciplinary approaches, combining various complementary techniques, especially those operating at a nanoscale level, potentially accelerate characterization of cancer cells or tissue properties. Here, we study a relation between the surface and biomechanical properties of melanoma cells, measured by mass spectrometry (ToF-SIMS) and atomic force microscopy (AFM). In total, seven cell lines have been studied. Six of them were melanoma cells derived from various stages of tumor progression: (1) WM115 cells derived from a 55 year old female skin melanoma at a vertical growth phase (VGP) in the primary melanoma site, (2) WM793 cells established from the vertical growth phase (VGP) of a primary skin melanoma lesion, (3) WM266-4 cells established from a cutaneous skin metastasis detected in the same patient as WM115 cells, (4) WM239 cells derived from a cutaneous skin metastasis, (5) 1205Lu cells originated from a lung metastasis diagnosed in the same patient as WM793 cells, and (6) A375P-cells were derived from a solid malignant tumor located in the lung. As a reference cell line, human epidermal melanocytes from adult skin (primary cell line HEMa-LP) were used. Results reveal low, medium, and large deformability of melanoma cells originating from vertical growth phase (VGP), and skin and lung metastasis, respectively. These changes were accompanied by distinct outcome from principal component analysis (PCA). In relation to VGP melanoma cells, cells from skin and lung metastasis reveal similar or significantly different surface properties. The largest deformability difference observed for cells from VGP and lung metastasis was accompanied by the largest separation of unspecific changes in their surface properties. In this way, we show the evidence that biomechanical and surface biochemical properties of cells change in parallel, indicating a potential of being used as nanobiophysical fingerprints of melanoma progression.

Influence of TiO2 Nanoparticles on Liquid Crystalline, Structural and Electrochemical Properties of (8Z)-N-(4-((Z)-(4-pentylphenylimino)methyl)benzylidene)-4-pentylbenzenamine.

Organic-inorganic hybrids based on liquid crystalline symmetrical imine (8Z)-N-(4-((Z)-(4-pentylphenylimino)methyl)benzylidene)-4-pentylbenzenamine (AZJ1) with two aliphatic chains and TiO2 nanomaterials were obtained and investigated taking into account thr crystallographic form of titanium dioxide i.e., anatase versus rutile. The type of TiO2 influences the mesomorphic properties of imine AZJ1, as observed by differential scanning calorimetry (DSC) and polarizing optical microscopy (POM) techniques. Fourier-Transform Infrared Spectroscopy (FT-IR) was used to investigate the interactions of oxygen vacancies located on the TiO2 surface with the studied AZJ1 imine together with studying the influence of temperature. Both imine:TiO2 anatase versus rutile hybrids possessed the highest occupied molecular orbital (HOMO) levels of about -5.39 eV (AZJ1:anatase) and -5.33 eV (AZJ1:rutile) and the lowest unoccupied molecular orbital (LUMO) levels of about -2.24 eV. The presence of TiO2 in each hybrid did not strongly affect the redox properties of imine AZJ1. Organic devices with the configuration of ITO/TiO2/AZJ1 (or AZJ1:TiO2 anatase versus rutile)/Au were fabricated and investigated in the presence and absence of visible light irradiation with a light intensity of 93 mW/cm². Finally, to analyze defects in the constructed organic devices we used thermal imaging and atomic force microscopy (AFM). The addition of TiO2 in both crystallographic forms has a positive influence on layer-forming properties that manifests itself as a very homogenous heat distribution for the whole sample.

Sequential binary protein patterning on surface domains of thermo-responsive polymer blends cast by horizontal-dipping.

We characterize an approach enabling dual protein positioning over broad polymer areas based on subsequent selective adsorption of two fluorescently labelled lectins, Concanavalin A (Con A) and Lentil Lectin (LcH), on self-assembled gradient patterns of thermoresponsive poly(N-isopropyl acrylamide) (PNIPAM) and polystyrene (PS) polymers blend, prepared by horizontal dipping technique. The film morphologies of gradient samples prior dual selective protein adsorption are mapped with scanning microscopy (AFM) and secondary ion mass spectrometry (ToF-SIMS), whereas adsorbed proteins are imaged with fluorescence microscope. ToF-SIMS analysis reveals surface composition consisting of PNIPAM-rich domains in PS-rich matrix. The two-step protein adsorption experiment results in selective adsorption of Con A and LcH to PNIPAM- and PS-rich phases, respectively. Integral geometry approach is used to compare quantitatively morphology of polymer patterns varied in domain size due to horizontal dipping casting. Minkowski measures are also used to compare quantitatively fluorescence micrographs of protein patches with SIMS images of original isotropic polymer patterns. It confirms that PNIPAM domains size increases with increasing speed. Further, Minkowski analysis unveiled that adsorbed proteins cover about 60-70% of polymer surface. What is more fluorescence micrographs acknowledge both no lectins contamination and no adsorption to interphase areas. Additionally, protein displacement effect is observed.

Between single ion magnets and macromolecules: a polymer/transition metal-based semi-solid solution.

The creation of functional magnetic materials for application in high-density memory storage or in the new field of molecular spintronics is a matter of widespread interest among the material research community. Herein, we describe a new approach that combines the qualities of single ion magnets, displaying slow magnetic relaxations, and the merits of polymers, being easy to process and widely used to produce thin films. Basing the idea on cobalt(ii) ions and pyridine-based single ion magnets, a new macromolecular magnetic material was obtained - a polymeric matrix of poly(4-vinylpyridine) (P4VP) cross-linked by a cobalt(ii) salt bound within it, effectively forming a network of single ion magnets, with field-induced magnetic relaxations preserved in both bulk and thin film forms. The binding of cobalt is confirmed by a series of methods, like secondary ion mass spectroscopy or high-resolution X-ray photoelectron spectroscopy. The magnetic relaxation times, up to 5 × 10-6 s, are controllable simply by dilution, making this new material a semi-solid solution. By this approach, a new path is formed to connect molecular magnetism and polymer science, showing that the easy polymer processing can be used in forming self-organizing functional magnetic thin films.

Synthesis and characterization of two new TiO2-containing benzothiazole-based imine composites for organic device applications.

The effect of the presence of titanium dioxide in two new imines, (E,E)-(butane-1,4-diyl)bis(oxybutane-4,1-diyl) bis(4-{[(benzo[d][1,3]thiazol-2-yl)methylidene]amino}benzoate) (SP1) and (E)-N-[(benzo[d][1,3]thiazol-2-yl)methylidene]-4-dodecylaniline (SP2), on the properties and stability of imine:TiO2 composites for organic device applications were examined. The investigated titanium dioxide (in anatase form, obtained via the sol-gel method) exhibited a surface area of 59.5 m2/g according to Brunauer-Emmett-Teller theory, and its structure is a combination of both meso- and microporous. The average pore diameter calculated by the Barrett-Joyner-Halenda method was 6.2 nm and the cumulative volume of pores was 0.117 m3/g. The imine SP1 exhibited columnar organization (Col), while SP2 revealed a hexagonal columnar crystalline phase (Colhk). The imine:TiO2 mixtures in various weight ratio (3:0, 3:1, 3:2, 3:3) showed a lower energy gap and HOMO-LUMO energy levels compared to pure TiO2. This implies that TiO2 provides not only a larger surface area for sensitizer adsorption and good electron collection, but also causes a shift of the imine energy levels resulting from intermolecular interaction. Also the temperature of the phase transition was slightly affected with the increase of TiO2 concentration in imine-based composites. The changes observed in the Fourier transform middle-infrared absorption (FT-MIR) spectra confirmed the significant influence of TiO2 on structural properties of both investigated imines. Similar interactions of oxygen vacancies existing on the TiO2 surface with SP1 and SP2 were observed. The imine:TiO2 mixtures showed good air stability and reusability, which demonstrates its potential for organic device applications.

Hybrid Materials Based on l,d-Poly(lactic acid) and Single-Walled Carbon Nanotubes as Flexible Substrate for Organic Devices.

We report on the application of l,d-poly(lactic acid) (l,d-PLA) with dispersed Single-Walled Carbon Nanotubes (SWCN) as a flexible translucent electrode for organic devices. We used commercially available nanotubes in various weight ratios from 0 to 8% dispersed in chloroform polymeric solution by ultrasonication and were drop cast. The created hybrid materials were investigated by differential scanning calorimetry to determine the influence of SWCN content on the thermal behavior, while polarizing optical microscope was used to find the effect of mechanical deformations on the textures. Drop-cast films were studied by optical transmittance, conductivity, dielectric properties and by thermal imaging under applied potential. Thermal imaging provided evidence of visible voltage-activated conduction. Simple mechanical deformation such as bending with stretching at edge to ca. 90 and elongation test were performed. Moreover, interactions between l,d-poly(lactic acid) and SWCN were investigated by FT-IR and NMR spectroscopy. Finally, we can conclude that the thermographic examination of created films permits fast, simple and inexpensive localization of defects on the surface of l,d-PLA:SWCN film, together with the electrical properties of the films.

Engineering a Poly(3,4-ethylenedioxythiophene):(Polystyrene Sulfonate) Surface Using Self-Assembling Molecules-A Chemical Library Approach.

The surface properties of poly(3,4-ethylenedioxythiophene):(polystyrene sulfonate) (PEDOT:PSS) affect the performance of many organic electronic devices. The work function determines the efficiency of the charge carrier transfer between PEDOT:PSS electrodes and the active layer of the device. The surface free energy affects phase separation in multicomponent blends that are typically used to fabricate active layers of organic light-emitting diodes and photovoltaic devices. Here, we present a method to prepare PEDOT:PSS films with a gradient work function and surface free energy. This modification was achieved by evaporation of trimethoxy(3,3,3-trifluoropropyl)silane in such a way that the degree of surface coverage of the molecules varied in the selected direction. Gradient films were used as electrodes to fabricate two-terminal PEDOT:PSS/poly(3-hexyl thiophene)/Au devices to rapidly screen for the influence of the modification on the performance of the prepared polymer diodes. Gradual changes in the morphology of the solution-cast model poly(3-butyl thiophene)/poly-bromostyrene films followed changes in the surface energy of the substrate.

Indirect immunoassay on functionalized silicon surface: Molecular arrangement, composition and orientation examined step-by-step with multi-technique and multivariate analysis.

The arrangement, composition and orientation of immunoreagents employed in an indirect immunoassay for determination of mycotoxin OchraToxin A (OTA) are specified for Si3N4 substrate, aiming to imitate biosensor transducers made of the same material. Si3N4 surfaces are examined after modification with (3-aminopropyl)triethoxysilane, spotting with OTA-ovalbumin conjugate (probe), blocking with bovine serum albumin, reaction with a mouse monoclonal antibody against OTA and, finally, reaction with a goat anti-mouse secondary antibody. Atomic force micrographs, their autocorrelation and height histogram parameters, show the stepwise development of a multi-component monolayer covered by groups of secondary antibody molecules. Time-Of-Flight Secondary Ion Mass Spectrometry reveals the composition of probe and blocking protein, as well as their partial desorption during the primary immunoreaction. Ellipsometry provides surface amount of all proteins, increasing step-by-step from 0.7 to 6.9mg/m2. In addition, ellipsometry combined with TOF-SIMS reveals the mass loadings of different molecules in the intermediate and the final overlayer. Based on this, some orientations of the immobilized molecules are proposed and a molar ratio of ∼2.5 for secondary to primary antibody is calculated. The orientations of the primary and secondary antibody are further clarified by Principal Component Analysis of TOF-SIMS data, through which a side-on and a head-on orientation is deduced for the primary and the secondary antibody, respectively. These findings demonstrate how the combination of multiple surface analysis techniques can provide insight on the arrangement, composition and orientation of biomolecules in the course of multi-step procedures employed in biosensors.

Orientation and biorecognition of immunoglobulin adsorbed on spin-cast poly(3-alkylthiophenes): Impact of polymer film crystallinity.

Many of bioelectronic and biosensor applications are based on poly(3-alkylthiophenes), conducting and solution-processable polymers. The most facile approach for the fabrication of such devices relies on biofunctionalization of P3AT surfaces with antibodies through adsorption. The success of this approach depends critically on antibody orientation that affects its biorecognition. As demonstrated here both these features are controlled by the surface structure of spin-cast P3ATs. In particular, a multi-technique and multivariate study that involved Atomic Force Microscopy, Grazing Incidence X-ray Diffraction, Angle-Resolved X-ray Photoelectron Spectroscopy, Enzyme-Linked ImmunoSorbent Assay, and Time-of-Flight Secondary Ion Mass Spectrometry combined with Principal Component Analysis is conducted in order to deduce the crystalline texture of three P3AT polymers as well as its effect on orientation of adsorbed rabbit immunoglobulin (IgG) molecules. An edge-on crystalline texture is concluded for regioregular poly(3-butylthiophene) (RP3BT) and poly(3-hexylthiophene) (RP3HT), while amorphous morphology is inferred for poly(3-butylthiophene) (P3BT). In addition, end-on and head-on orientations similar for all P3ATs were concluded, based on the amount of adsorbed rabbit IgG molecules. Examination of amino acids characteristic for F(ab')2 and Fc fragments, and dominant in the external regions of adsorbed immunoglobulin molecules, points to end-on IgG alignment on RP3BT and RP3HT, but not on P3BT. Moreover, the binding of an anti-rabbit IgG antibody on the absorbed rabbit IgG is higher (up to 71%) when the biorecognition reactions are performed on regioregular rather than regiorandom P3AT surfaces. In particular, the highest biorecognition efficiency and IgG orientational order is observed for the RP3BT surfaces with the more developed crystallinity.

Protocol of single cells preparation for time of flight secondary ion mass spectrometry.

There are several techniques like time of flight secondary ion mass spectrometry (ToF SIMS) that require a special protocol for preparation of biological samples, in particular, those containing single cells due to high vacuum conditions that must be kept during the experiment. Frequently, preparation methodology involves liquid nitrogen freezing what is not always convenient. In our studies, we propose and validate a protocol for preparation of single cells. It consists of four steps: (i) paraformaldehyde fixation, (ii) salt removal, (iii) dehydrating, and (iv) sample drying under ambient conditions. The protocol was applied to samples with single melanoma cells i.e. WM115 and WM266-4 characterized by similar morphology. The surface and internal structures of cells were monitored using atomic force, scanning electron and fluorescent microscopes, used to follow any potential protocol-induced alterations. To validate the proposed methodology for sample preparation, ToF SIMS experiments were carried out using C60(+) cluster ion beam. The applied principal component analysis (PCA) revealed that chemical changes on cell surface of melanoma cells were large enough to differentiate between primary and secondary tumor sites. Subject category: Mass spectrometry.

Thiolate versus Selenolate: Structure, Stability, and Charge Transfer Properties.

Selenolate is considered as an alternative to thiolate to serve as a headgroup mediating the formation of self-assembled monolayers (SAMs) on coinage metal substrates. There are, however, ongoing vivid discussions regarding the advantages and disadvantages of these anchor groups, regarding, in particular, the energetics of the headgroup-substrate interface and their efficiency in terms of charge transport/transfer. Here we introduce a well-defined model system of 6-cyanonaphthalene-2-thiolate and -selenolate SAMs on Au(111) to resolve these controversies. The exact structural arrangements in both types of SAMs are somewhat different, suggesting a better SAM-building ability in the case of selenolates. At the same time, both types of SAMs have similar packing densities and molecular orientations. This permitted reliable competitive exchange and ion-beam-induced desorption experiments which provided unequivocal evidence for a stronger bonding of selenolates to the substrate as compared to the thiolates. Regardless of this difference, the dynamic charge transfer properties of the thiolate- and selenolate-based adsorbates were found to be nearly identical, as determined by the core-hole-clock approach, which is explained by a redistribution of electron density along the molecular framework, compensating the difference in the substrate-headgroup bond strength.

Differentiation between single bladder cancer cells using principal component analysis of time-of-flight secondary ion mass spectrometry.

Time-of-flight-secondary ion mass spectrometry (TOF-SIMS) mass spectra measurements combined with an appropriate sample preparation protocol are the powerful tools to obtain unique information about the chemical composition of biological materials. In our studies, two questions were addressed, i.e., whether it is possible to develop a fixative-based sample preparation protocol and whether it allows one to distinguish between cells originating from various stages of cancer progression. Therefore, four human bladder cancer cell lines (with distinct malignancy degree) have been investigated. A chemical fixation protocol has been used for TOF-SIMS measurements, and mass spectra were obtained using a Bi3(+) primary ion beam. The principal component analysis (PCA) has been applied to analyze the whole range of mass spectra (without preselection of any particular masses) using two approaches of data preprocessing, namely, mean centering and autoscaling. The PC3 versus PC2 plot has showed significant differences between nonmalignant cancer cells and the cancerous ones for both of preprocessing approaches. The analysis of mass spectra of human bladder cells allows one to find a list of mass peaks with intensities significantly larger in cancerous bladder cells compared to nonmalignant cell cancer of the ureter (HCV29 cells). These findings show that TOF-SIMS in combination with PCA can be used to identify reference, human bladder cells from cancerous ones.

Imaging and spectroscopic comparison of multi-step methods to form DNA arrays based on the biotin-streptavidin system.

Three multi-step multi-molecular approaches using the biotin-streptavidin system to contact-print DNA arrays on SiO2 surfaces modified with (3-glycidoxypropyl)trimethoxysilane are examined after each deposition/reaction step by atomic force microscopy, X-ray photoelectron spectroscopy and time of flight secondary ion mass spectrometry. Surface modification involves the spotting of preformed conjugates of biotinylated oligonucleotides with streptavidin onto surfaces coated with biotinylated bovine serum albumin b-BSA (approach I) or the spotting of biotinylated oligonucleotides onto a streptavidin coating, the latter prepared through a reaction with immobilized b-BSA (approach II) or direct adsorption (approach III). AFM micrographs, quantified by autocorrelation and height histogram parameters (e.g. roughness), reveal uniform coverage after each modification step with distinct nanostructures after the reaction of biotinylated BSA with streptavidin or of a streptavidin conjugate with biotinylated oligonucleotides. XPS relates the immobilization of biomolecules with covalent binding to the epoxy-silanized surface. Protein coverage, estimated from photoelectron attenuation, shows that regarding streptavidin the highest and the lowest immobilization efficiency is achieved by following approaches I and III, respectively, as confirmed by TOF-SIMS microanalysis. The size of the DNA spot reflects the contact radius of the printed droplet and increases with protein coverage (and roughness) prior to the spotting, as epoxy-silanized surfaces are hardly hydrophilic. Representative TOF-SIMS images show sub-millimeter spots: uniform for approach I, doughnut-like (with a small non-zero minimum) for approach II, both with coffee-rings or peak-shaped for approach III. Spot features, originating from pinned contact lines and DNA surface binding and revealed by complementary molecular distributions (all material, DNA, streptavidin, BSA, epoxy, SiO2), indicate two modes of droplet evaporation depending on the details of each applied approach.