Wolfgang Fritzsche

Mapping elasticity of rehydrated metaphase chromosomes by scanning force microscopy

Scanning force microscopy was used for mapping the viscoelastic properties of metaphase chromosomes. These properties were probed by scanning with various imaging forces and subsequent calculation of the difference image. The procedure allows a mapping of the viscoelastic behavior expressed as force-dependent indentation of the local surface feature and results in an image with material contrast. The approach is demonstrated on rehydrated metaphase chromosomes, which were spread and air-dried before rehydration in aqueous buffer. The rehydration resulted in a swelling of the chromosome structure and was accompanied by drastic changes in the viscoelastic properties. For comparisons, force-distance curves on metaphase chromosomes were accumulated; these curves were also used for the calculation of the stiffness curve. The demonstrated approach of mapping viscoelasticity by differential scanning force microscopy allows the detection of domains with varying mechanical properties in biomolecules such as chromosomes.

Scanning force microscopy reveals ellipsoid shape of chicken erythrocyte nucleosomes

Scanning force microscopy was used to investigate the conformation of hypotonic spread chicken erythrocyte nucleosomes. Nucleosomal chains were prepared in low-salt conditions and fixed before centrifugation onto glass coverslips and air drying. The images of single nucleosomes were isolated by image processing, and the height and geometry of the resulting three-dimensional structures were investigated. An average nucleosome height of 4.2 +/- 1.1 nm was determined. A virtual cross section at half-maximum height of the nucleosome structure was used for a characterization of the nucleosome geometry. The shape of this cross section was best described by an ellipse with an aspect ratio (major/minor axis) of approximately 1.30.

Nanoparticle-based optical detection of molecular interactions of DNA-chip technology

We adapted the nanoparticle-labeling technique from microscopical applications for DNA-chip detection. Nanoparticles can be detected by simple optical means, and exhibit a high stability. So alternative optical readout devices can be applied for the detection of specific DNA- binding on microstructured spots of complementary, surface- immobilized capture DNA. Several devices were tested, and the binding of gold-labeled DNA to complementary and non- complementary sequences was investigated using optical detection of the gold-labeled substrates before and after silver enhancement.

Ultrastructural characterization of colloidal metal films for bioanalytical applications by scanning force microscopy

Colloidal metal films (CMFs) are prepared by the attachment of silver, gold, or platinum (or other metal) particles to a glass slide modified by silanization with 3‐mercaptopropyl silane. The covalent attachment of the metal particles occurs through the metal–sulfur bond. In these samples the local electromagnetic field is enhanced near the surface of the CMF due to excitation of plasmon resonances. This phenomenon can be used for a variety of analytical applications. Because the optical properties are strongly dependent on the morphology of the film, its structural characterization becomes of great importance. To further characterize CMFs we have used scanning force microscopy (SFM). Initial studies revealed lateral dimensions of the particles as well as the particle density. Height measurements were made using the three‐dimensional topographic image of the surface yielded by SFM, and were used to evaluate the selective deposition of a silica spacer layer onto the metal particles. Comparative SFM measure...

Metal Nanoparticles for Molecular Plasmonics

This review provides an overview about nanoparticle synthesis and the characterization of related optical properties of homogeneous as well as bimetallic nanoparticles made of gold, silver, and other metals. The optical properties of metal nanoparticles (plasmonic nanoparticles) depend strongly on the metal that is used, but also on the size, the shape, the surrounding medium, and the interparticle distance. The progress in the controlled synthesis as well as sophisticated bioconjugation of metal nanoparticles in combination with molecular principles lead to the new field of nanoparticle-based molecular nanotechnology, the molecular plasmonics. Applications are envisioned in the fields of bioanalytics and nanooptics.

Reconstruction of ribosomal subunits and rDNA chromatin imaged by scanning force microscopy

Scanning force microscopy (SFM) reveals surface topography by scanning a sharp tip in close proximity to the sample. Due to tip–sample interaction, artificial broadening of the real surface structure with the tip geometry occurs. One approach for image reconstruction is the use of calibration standards, preferably in the size range of the samples. In the present study, an image reconstruction method based on colloidal gold as a geometric standard was used to reconstruct SFM images of biomolecules. Sample and calibration standard size were in the nanometer range, and the standards were coadsorbed with the specimen. Raw and reconstructed images of the biomolecules were compared, and the reconstruction was characterized by difference images as well as determination of the difference volume. The application of image reconstruction based on colloidal gold as a calibration standard for SFM of biomolecules is discussed.

Optical Detection Of DNA Constructs Based On Nanoparticles And Silver Enhancement

Nanoparticle‐labeling was recently introduced for probing immobilized DNA by scanning force microscopy or optical detection. The optical detection has the potential of high parallelization in combination with miniaturization, thereby enabling a high sample throughput. However, a quantification of the optical signal and a correlation of this signal with the surface density of bound nanoparticles are needed. We will demonstrate the application of the silver enhancement procedure for a signal amplification to extend the dynamic range of the method. The specificity of the enhanced nanoparticle‐labeling will be shown, and the influence of the surface density of immobilized molecules on the signal is studied. The results confirm that the proposed detection scheme is suitable for an application in molecular nanotechnology for the characterization of DNA‐modified surfaces.

Analysis of G‐5x Quadruplex DNA

Pure guanine‐rich oligonucleotide (GRO) DNA will self‐assemble into quadruplex “G‐wire” DNA in the absence of stabilizing monovalent cations. Atomic force microscopy of GRO G‐wires bound to freshly cleaved mica and imaged in air reveal extensive networks of connected quadruplex DNA auto‐oriented on the next nearest neighbor spacing of the mica substrate. Electrical measurements of GRO G‐wires across micro‐fabricated gold electrodes on silicon oxide indicate the GRO G‐wire networks are electrical insulators. The suspected random nature of the GRO incorporation into quadruplex DNA suggested an alternative mechanism of incorporating metal ions into G‐wires: through direct weak ionic binding to the extensive phosphate backbone exposure of both the quadruplex and partially assembled oligonucelotide sequences. High densities of GRO mixed with CuCl2 or AgNO3 produced dramatically wider wires that we believe are combinations of weakly interacting metal ions and GRO. After reduction by exposure to light the Ag+ do...

Printed circuit boards as platform for disposable lab-on-a-chip applications

An increasing demand in performance from electronic devices has resulted in continuous shrinking of electronic components. This shrinkage has demanded that the primary integration platform, the printed circuit board (PCB), follow this same trend. Today, PCB companies offer ~100 micron sized features (depth and width) which mean they are becoming suitable as physical platforms for Lab-on-a-Chip (LOC) and microfluidic applications. Compared to current lithographic based fluidic approaches; PCB technology offers several advantages that are useful for this technology. These include: Being easily designed and changed using free software, robust structures that can often be reused, chip layouts that can be ordered from commercial PCB suppliers at very low cost (1 AUD each in this work), and integration of electrodes at no additional cost. Here we present the application of PCB technology in connection with microfluidics for several biomedical applications. In case of commercialization the costs for each device can be even further decreased to approximately one tenth of its current cost.

Coupling G‐Wires To Metal Nanoparticles

G‐wires are DNA superstructures based on quartet formation by four guanine (G) bases. These molecular structures reach the micrometer size scale by assembling short oligonucleotides of a guanine rich sequence. Physico‐chemical parameters were optimized to achieve optimal superstructure assembly. The combination of these superstructures with metal nanoparticles could provide new tools for a molecular nanotechnology. An assembly strategy has been developed and first experiments towards an inclusion of nanoparticles in the supramolecular assembly were conducted.