Mechanische Eigenschaften von freitragenden artifiziellen und nativen Membranen

This thesis presents a new method to explore the local mechanical properties such as bending modulus or surface tension of artificial and native pore-spanning membranes. Therefore the elastic response of a free-standing membrane to a local indentation by the means of atomic force microscopy is measured. Starting point are highly hexagonal ordered pores in alumina produced by electrochemical anodization of planar aluminium. The homogeneous pore radius can by tailored in the range of 10 nm up to 200 nm, but radius of 33 nm, 90 nm and 200 nm turned out to be best suited for investigation of the mechanical properties of pore-spanning native or artificial membranes. In this work artificial membrane systems consisting of DODAB as a bilayer in gel phase or DOTAP as a fluide membrane are spreaded by vesicle absorption on hexagonal structured pores after chemisorption of a 3-mercaptopropionic acid monolayer. Centrally indenting these nanodrums with an atomic force microscope tip yields force-indentation curves, which are quantitatively analyzed by solving the corresponding shape equations of continuum curvature elasticity. Since the measured response depends in a known way on the system geometry (pore size, tip radius) and on material parameters (bending modulus, lateral tension, adhesion), this opens the possibility to monitor local elastic properties of lipid membranes in a well-controlled setting. Additionally the locally distributed mechanical properties of pore-spanning artificial membranes are compared to those of native pore-spanning membranes. Therefore the basal membrane of MDCK II cells was prepared on porous alumina assays and their mechanical properties were analyzed by means of atomic force microscopy. Finally the elastic behavior such as the Young modulus of living MDCK II cells under various osmotic pressures is investigated. By changing the osmolarity in the extracellular region of MDCK II cells a volume change is induced according to hydration and dehydration of the cells, respectively. This volume change induces also a change in the elastic behavior of the cell, which is quantified by the means of force spectroscopy

Nanostructured cavity devices for extracellular stimulation of HL-1 cells

Microelectrode arrays (MEAs) are state-of-the-art devices for extracellular recording and stimulation on biological tissue. Furthermore, they are a relevant tool for the development of biomedical applications like retina, cochlear and motor prostheses, cardiac pacemakers and drug screening. Hence, research on functional cell-sensor interfaces, as well as the development of new surface structures and modifications for improved electrode characteristics, is a vivid and well established field. However, combining single-cell resolution with sufficient signal coupling remains challenging due to poor cell-electrode sealing. Furthermore, electrodes with diameters below 20 µm often suffer from a high electrical impedance affecting the noise during voltage recordings. In this study, we report on a nanocavity sensor array for voltage-controlled stimulation and extracellular action potential recordings on cellular networks. Nanocavity devices combine the advantages of low-impedance electrodes with small cell-chip interfaces, preserving a high spatial resolution for recording and stimulation. A reservoir between opening aperture and electrode is provided, allowing the cell to access the structure for a tight cell-sensor sealing. We present the well-controlled fabrication process and the effect of cavity formation and electrode patterning on the sensor's impedance. Further, we demonstrate reliable voltage-controlled stimulation using nanostructured cavity devices by capturing the pacemaker of an HL-1 cell network

A Label-Free Optical Detection of Pathogens in Isopropanol as a First Step towards Real-Time Infection Prevention

The detection of pathogens is a major public health issue. Every year, thousands of people die because of nosocomial infections. It is therefore important to be able to detect possible outbreaks as early as possible, especially in the hospital environment. Various pathogen detection techniques have already been demonstrated. However, most of them require expensive and specific equipment, and/or complex protocols, which, most of the time, involve biochemical reaction and labelling steps. In this paper, a new method that combines microscopic imaging and machine learning is described. The main benefits of this approach are to be low-cost, label-free and easy to integrate in any suitable medical device, such as hand hygiene dispensers. The suitability of this pathogen detection method is validated using four bacteria, both in PBS (Phosphate Buffered Saline) and in isopropanol. In particular, we demonstrated an efficient pathogenic detection that is sensible to changes in the composition of a mixture of pathogens, even in alcohol-based solutions

A fluorescence based method for the quantification of surface functional groups in closed micro- and nanofluidic channels

Surface analysis is critical for the validation of microfluidic surface modifications for biology, chemistry, and physics applications. However, until now quantitative analytical methods have mostly been focused on open surfaces. Here, we present a new fluorescence imaging method to directly measure the surface coverage of functional groups inside assembled microchannels over a wide dynamic range. A key advance of our work is the elimination of self-quenching to obtain a linear signal even with a high density of functional groups. This method is applied to image the density and monitor the stability of vapor deposited silane layers in bonded silicon/glass micro- and nanochannels

Knowledge about Hand Hygiene and Related Infectious Disease Awareness among Primary School Children in Germany †

Hand hygiene is a cornerstone of infection prevention. However, few data are available for school children on their knowledge of infectious diseases and their prevention. The aim of the study was to develop and apply a standardized questionnaire for children when visiting primary schools to survey their knowledge about infectious diseases, pathogen transmission and prevention measures. Enrolling thirteen German primary schools, 493 questionnaires for grade three primary school children were included for further analyses, comprising 257 (52.1%) girls and 236 (47.9%) boys with an age range of 8–11 years. Out of 489 children, 91.2% participants indicated that they knew about human-to-human transmissible diseases. Of these, 445 children responded in detail, most frequently mentioning respiratory and gastrointestinal diseases, followed by childhood diseases. Addressing putative hygiene awareness-influencing factors, it was worrisome that more than 40.0% of the children avoided visiting the sanitary facilities at school. Most of the children (82.9%) noted that they did not like to use the sanitary facilities at school because of their uncleanliness and the poor hygienic behavior of their classmates. In conclusion, basic infection awareness exists already in primary school age children. Ideas about the origin and prevention of infections are retrievable, however, this knowledge is not always accurate and adequately contextualized. Since the condition of sanitary facilities has a strong influence on usage behavior, the child’s perspective should be given more consideration in the design and maintenance of sanitary facilities

Mechanical Properties of Pore-Spanning Lipid Bilayers Probed by Atomic Force Microscopy

We measure the elastic response of a free-standing lipid membrane to a local indentation by using an atomic force microscope. Starting point is a planar gold-coated alumina substrate with a chemisorbed 3-mercaptopropionic acid monolayer displaying circular pores of very well defined and tunable size, over which bilayers composed of N,N,-dimethyl-N,N,- dioctadecylammonium bromide or 1,2-dioleoyl-3-trimethylammonium-propane chloride were spread. Centrally indenting these ‘‘nanodrums’’ with an atomic force microscope tip yields force-indentation curves, which we quantitatively analyze by solving the corresponding shape equations of continuum curvature elasticity. Since the measured response depends in a known way on the system geometry (pore size, tip radius) and on material parameters (bending modulus, lateral tension), this opens the possibility to monitor local elastic properties of lipid membranes in a well-controlled setting.</p

Practical aspects of Boersch phase contrast electron microscopy of biological specimens

Walter A, Muzik H, Vieker H, et al. Practical aspects of Boersch phase contrast electron microscopy of biological specimens. Ultramicroscopy. 2012;116:62-72.Implementation of physical phase plates into transmission electron microscopes to achieve in-focus contrast for ice-embedded biological specimens poses several technological challenges. During the last decade several phase plates designs have been introduced and tested for electron ciyo-microscopy (cryoEM), including thin film (Zernike) phase plates and electrostatic devices. Boersch phase plates (BPPs) are electrostatic einzel lenses shifting the phase of the unscattered beam by an arbitrary angle. Adjusting the phase shift to 90 achieves the maximum contrast transfer for phase objects such as biomolecules. Recently, we reported the implementation of a BPP into a dedicated phase contrast aberration-corrected electron microscope (PACEM) and demonstrated its use to generate in-focus contrast of frozen-hydrated specimens. However, a number of obstacles need to be overcome before BPPs can be used routinely, mostly related to the phase plate devices themselves. CryoEM with a physical phase plate is affected by electrostatic charging, obliteration of low spatial frequencies, and mechanical drift. Furthermore, BPPs introduce single sideband contrast (SSB), due to the obstruction of Friedel mates in the diffraction pattern. In this study we address the technical obstacles in detail and show how they may be overcome. We use X-ray photoelectron spectroscopy (XPS) and Auger electron spectroscopy (AES) to identify contaminants responsible for electrostatic charging, which occurs with most phase plates. We demonstrate that obstruction of low-resolution features is significantly reduced by lowering the acceleration voltage of the microscope. Finally, we present computational approaches to correct BPP images for SSB contrast and to compensate for mechanical drift of the BPP. (C) 2012 Elsevier B.V. All rights reserved