Silvia Mittler

Sensitivity studies for specific binding reactions using the biotin/streptavidin system by evanescent optical methods

The combination of various evanescent optical methods such as surface plasmon spectroscopy, waveguide mode spectroscopy and an integrated optical Mach-Zehnder-interferometer are used to characterize biotinylated self-assembled monolayers as well as the binding of streptavidin to these labels. The aim of designing a highly specific and sensitive, re-usable affinity sensor for antigens on the basis of an integrated optical Mach-Zehnder interferometer is based on a proper understanding of the characteristics of the entire binding matrix architecture. Therefore, a variety of biotin-derivatives immobilized in a monolayer are investigated with respect to their affinity to streptavidin and the possibility to remove the steptavidin layer specifically. The density of the streptavidin layer as well as the optical constants of the involved molecules are measured. Finally the integrated optical Mach-Zehnder interferometer is tested with respect to the sensitivity to an antigen-antibody binding reaction. An attempt to further increase the sensitivity by simultaneous detection of a fluorescence signal failed due to bleaching effects.

Disulfide-Linked, Gold Nanoparticle Based Reagent for Detecting Small Molecular Weight Thiols

Disulfide-linked gold nanoparticles (AuNP) were synthesized by reacting dithiobis[succinimidylpropionate] (DSP) coated nanoparticles with glutathione disulfide. AuNP-cross-linking was monitored by the red shift and broadening of the AuNPs localized surface plasmon absorption resonance (LSPR) spectrum. The exposure of the disulfide-linked AuNPs to a variety of free thiols with systematically varying molecular weight revealed a AuNP-disulfide stability to reduction by thiols up to a critical molecular weight, M(c), of >300 Da thus making the disulfide-linked AuNP the first reagent that can discriminate thiols based on their size.

Migration of Periodontal Ligament Fibroblasts on Nanometric Topographical Patterns: Influence of Filopodia and Focal Adhesions on Contact Guidance

Considered to be the “holy grail” of dentistry, regeneration of the periodontal ligament in humans remains a major clinical problem. Removal of bacterial biofilms is commonly achieved using EDTA gels or lasers. One side effect of these treatment regimens is the etching of nanotopographies on the surface of the tooth. However, the response of periodontal ligament fibroblasts to such features has received very little attention. Using laser interference lithography, we fabricated precisely defined topographies with continuous or discontinuous nanogrooves to assess the adhesion, spreading and migration of PDL fibroblasts. PDL fibroblasts adhered to and spread on all tested surfaces, with initial spreading and focal adhesion formation slower on discontinuous nanogrooves. Cells had a significantly smaller planar area on both continuous and discontinuous nanogrooves in comparison with cells on non-patterned controls. At 24 h post seeding, cells on both types of nanogrooves were highly elongated parallel to the groove long axis. Time-lapse video microscopy revealed that PDL fibroblast movement was guided on both types of grooves, but migration velocity was not significantly different from cells cultured on non-patterned controls. Analysis of filopodia formation using time-lapse video microscopy and labeling of vinculin and F-actin revealed that on nanogrooves, filopodia were highly aligned at both ends of the cell, but with increasing time filopodia and membrane protrusions developed at the side of the cell perpendicular to the cell long axis. We conclude that periodontal ligament fibroblasts are sensitive to nanotopographical depths of 85–100 µm, which could be utilized in regeneration of the periodontal ligament.

Gold and thiol surface functionalized integrated optical Mach–Zehnder interferometer for sensing purposes

Abstract We demonstrate an integrated optical Mach–Zehnder interferometer for sensing purposes with a surface functionalized by thiols via an additional gold layer. Therefore, the oxidic silicon oxynitride waveguide surface was first covered by a thiol terminated silane self-assembled monolayer. An ultrathin gold layer was then deposited onto the “active” sulfur surface by OMCVD. This gold layer was finally functionalized by a biotinylated thiol in order to specifically bind streptavidin and a biotinylated antibody. Unspecific binding of streptavidin was tested also.

An integrated optical Mach–Zehnder interferometer functionalized by β-cyclodextrin to monitor binding reactions

Abstract We demonstrate an integrated optical Mach–Zehnder interferometer for the monitoring of guest–host reactions on immobilized β-cyclodextrin (β-CD). Adamantanecarboxylic acid, 4-tert-butylbenzoic acid and methyl orange were investigated as guest molecules. The binding constants of the three guests into the two-dimensionally arranged immobilized cyclodextrins could be determined and are a quarter to a third smaller than the values measured in the three-dimensional environment of a solution.

Waveguide evanescent field fluorescence microscopy: Thin film fluorescence intensities and its application in cell biology

We demonstrate an inexpensive alternative to total internal reflection fluorescence microscopy. A method for imaging ultrathin films and living cells located on waveguides—illuminated with their evanescent fields—is introduced. An extensive analysis of ion-exchanged waveguides focusing on their application as microscopy substrates for studying interfacial phenomena is presented. Experimental results are in excellent agreement with the simulations. As an application osteoblasts (bone matrix forming cells) and ultrathin Langmuir–Blodgett films were imaged. The fluorescence intensity has been used to determine the cell attachment.

Selective Nucleation and Area-Selective OMCVD of Gold on Patterned Self-Assembled Organic Monolayers Studied by AFM and XPS: A Comparison of OMCVD and PVD

We demonstrate the area selective organometallic chemical vapor deposition (OMCVD) of ultrathin gold layers, using (trimethylphosphine)methylgold [(CH 3 ) 3 P]AuCH 3 as the gold source, onto self-assembled monolayers (SAMs) of ω-functionalized, long chain alkanethiols on gold and silver coated silicon and mica samples. The dependence on the reaction time of surface coverage with gold nano-clusters is analyzed by atomic force microscopy (AFM). X-ray photoelectron spectroscopy (XPS) shows that nucleation and growth occur only on thiol functionalized surfaces and not on methyl or hydroxyl functionalized SAMs. The selectivity of the growth is completely lost if gold is deposited by thermal evaporation of the pure metal, as shown in direct comparison with the OMCVD method employing mixed SAMs of different surface reactivity (-SH vs. -CH 3 ) that were patterned by microcontact printing.

Digital in-line soft x-ray holography with element contrast

Digital in-line soft x-ray holography (DIXH) was used to image immobilized polystyrene and iron oxide particles and to distinguish them based on their different x-ray absorption cross sections in the vicinity of the carbon K-absorption edge. The element-specific information from the resonant DIXH images was correlated with high-resolution scanning electron microscopy (SEM) pictures. We also present DIXH images of a cell nucleus and compare the contrast obtained for nuclear components with the appearance in optical microscopy.

3-D FDTD Analysis of Gold-Nanoparticle-Based Photonic Crystal on Slab Waveguide

In this paper, metallic photonic crystals (PC) based on 2-D periodic arrays of gold nanoparticles were investigated on indium tin oxide slab waveguides using 3-D finite-difference time domain simulations with nonuniform mesh techniques. The PC effects were studied by changing the lattice constants from 300 to 500 nm. The results obtained indicate that the waveguide-excited plasmon absorption peak of periodic array of gold nanoparticles is tunable from 672 to 707 nm due to the second grating order propagating backward at the grazing angle. The nanoparticle-induced extinction of the waveguide mode was also investigated by varying the slab thickness from 100 to 375 nm. The results show that the extinction peak shifts from 650 to 705 nm. The theoretical results predict that the interactions of the periodic array of gold nanoparticles are strongly affected by the dispersion of the waveguide.

Functional polymers as nanoscopic building blocks

Abstract Polyphenylene dendrimers are introduced as polymeric building blocks—with a strictly monodisperse particle size distribution within the nanometer range—for the construction of nanostructured materials and devices. The possibility for the introduction of different functionalities in the core, the scaffold or the periphery of the dendrimers offer their use as interesting modules for photonic, electronic or bioactive structures and supramolecular functional assemblies. Thus, dendrimers complement the available set of nanoscopic building blocks made from metals, e.g., Au nanoclusters and semiconductors, e.g., luminescent quantum dots. In a first set of experiments, we describe the fabrication of multilayer architectures using dendrimers with chargeable groups at the surface. This way, the polyelectrolyte deposition technique can be applied for the construction of hybrid layered assemblies with a control of the internal supramolecular structure at the nanometer level. Surface plasmon field-enhanced fluorescence spectroscopy is used to monitor the luminescent properties of dendrimers with a phthalocyanine core integrated into such a multilayer assembly. AFM and SEM micrographs demonstrate the use of surface-functionalized dendrimers (exposing sulfur groups at the periphery) in combination with Au nanoparticles for the controlled assembly of hybrid aggregates as nanoscopic functional devices.