Reinhard Guckenberger

Inverting dynamic force microscopy: From signals to time-resolved interaction forces

Transient forces between nanoscale objects on surfaces govern friction, viscous flow, and plastic deformation, occur during manipulation of matter, or mediate the local wetting behavior of thin films. To resolve transient forces on the (sub) microsecond time and nanometer length scale, dynamic atomic force microscopy (AFM) offers largely unexploited potential. Full spectral analysis of the AFM signal completes dynamic AFM. Inverting the signal formation process, we measure the time course of the force effective at the sensing tip. This approach yields rich insight into processes at the tip and dispenses with a priori assumptions about the interaction, as it relies solely on measured data. Force measurements on silicon under ambient conditions demonstrate the distinct signature of the interaction and reveal that peak forces exceeding 200 nN are applied to the sample in a typical imaging situation. These forces are 2 orders of magnitude higher than those in covalent bonds.

Enhancing the resolution of scanning near-field optical microscopy by a metal tip grown on an aperture probe

We show improvement of the optical and topographical resolution of scanning near-field optical microscopy by introducing a “tip-on-aperture” probe, a metallic tip formed on the aperture of a conventional fiber probe. The tip concentrates the light passing through the aperture. Thus the advantages of aperture and apertureless scanning near-field optical microscopy are combined. Tips are grown by electron beam deposition and then covered with metal. Fluorescent beads are imaged with a resolution down to 25 nm (full width at half maximum) in the optical signal. The near-field appears strongly localized within 5 nm in z direction, thus promising even higher resolution with sharper tips.

From Images to Interactions: High-Resolution Phase Imaging in Tapping-Mode Atomic Force Microscopy

In tapping-mode atomic force microscopy, the phase shift between excitation and response of the cantilever is used as a material-dependent signal complementary to topography. The localization of information in the phase signal is demonstrated with 1.4-nm lateral resolution on purple membrane of Halobacterium salinarum in buffer solution. In a first-order approximation, the phase signal is found to correlate with modulations of the tip oscillation amplitude, induced by topography. Extending the analysis to contributions of the tip-sample interaction area as a second-order approximation, a method is proposed to extract information about the interaction from the phase signal for surfaces with a roughness in the order of the tip radius.

Higher-harmonics generation in tapping-mode atomic-force microscopy: Insights into the tip–sample interaction

We present an experimental analysis of the nonlinear tip–sample interaction in tapping-mode atomic-force microscopy by exploiting anharmonic contributions of the cantilever motion. Two aspects of a concept aiming at a full reconstruction of the tip–sample interaction are demonstrated: higher flexural eigenmode vibrations excited by the impact of the oscillating tip on the sample are used to measure the tip–sample interaction time; by imaging at higher harmonics of the driving frequency material contrast is obtained.

Determination of a common origin in the micrographs of tilt series in three-dimensional electron microscopy

Abstract In 3D electron microscopy a common origin in micrographs of a tilt series is usually determined by means of correlation functions. Until now proof of the validity of this method has only been provided for an object consisting of points. This paper proves the method to be valid in the general case of a continuous object imaged linearly and discusses some theoretical and practical aspects.

Contrast of microwave near-field microscopy

Constant-height scanning is demonstrated to improve near-field microscopy by eliminating artifacts connected with topography scanning, hence, to image the inherent electromagnetic contrast. Microwaves are chosen for this study because the long wavelength eliminates coherence artifacts, owing to a scale separation of wave and image frequencies. Measured amplitude and phase images of conductive films are quantitatively analyzed by considering the longitudinal electric near field. The observed spatial resolution of 200 nm equals the probing tip size and proves that the skin depth δ of the tip material (here, 1600 nm) presents no resolution limit to scanning optical microscopy.

Distribution of the surfactant-associated protein C within a lung surfactant model film investigated by near-field optical microscopy.

Lung surfactant films at the air/water interface exhibit the particularity that surfactant molecules are expelled from the surface monolayer into a surface associated multilamellar phase during compression. They are able to re-enter the surface film during the following expansion. The underlying mechanism for this behavior is not fully understood yet. However, an important role is ascribed to the surfactant-associated protein C (SP-C). Here, we studied a model lung surfactant, consisting of dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylglycerol (DPPG), and SP-C, by means of scanning near-field optical microscopy (SNOM). Attaching a fluorescent dye to the protein allowed the localization of its lateral distribution at various surface pressures with high resolution. At an early stage of compression, the film appears demixed into a pure lipid phase and a protein-enriched phase. Within the latter phase, protein aggregations are revealed. They show a uniform density, having three times the fluorescence intensity of their surroundings. Across the phase boundary between the lipid phase and the protein-rich phase, there is a protein density gradient rather than an abrupt border. When the film is highly compressed, we observe the formation of multilamellar structures that are fluorescent. They are often surrounded by a slightly fluorescent monolayer. The fluorescence of the multilayer stacks (i. e., the protein content per unit area) is proportional to the height of the stacks.

Scanning Tunneling Microscopy of a Hydrated Bacterial Surface Protein

Abstract Imaging of biological macromolecules by scanning tunneling microscopy is hampered by the poor electrical conductivity of these objects. We found, however, that by controlling the protein hydration a small but sufficient conductivity can be induced. At tunneling currents lower than 0.5 pA and tunneling voltages up to 10 V and at a humidity in the range between 30% and 45% it was possible to obtain clear images of the HPI layer, a protein which naturally forms a two-dimensional ordered array on a bacterial surface.

Native protein nanolithography that can write, read and erase

The development of systematic approaches to explore protein-protein interactions and dynamic protein networks is at the forefront of biological sciences. Nanopatterned protein arrays offer significant advantages for sensing applications, including short diffusion times, parallel detection of multiple targets and the requirement for only tiny amounts of sample. Atomic force microscopy (AFM) based techniques have successfully demonstrated patterning of molecules, including stable proteins, with submicrometre resolution. Here, we introduce native protein nanolithography for the nanostructured assembly of even fragile proteins or multiprotein complexes under native conditions. Immobilized proteins are detached by a novel vibrational AFM mode (contact oscillation mode) and replaced by other proteins, which are selectively self-assembled from the bulk. This nanolithography permits rapid writing, reading and erasing of protein arrays in a versatile manner. Functional protein complexes may be assembled with uniform orientation at dimensions down to 50 nm. Such fabrication of two-dimensionally arranged nano-objects with biological activity will prove powerful for proteome-wide interaction screens and single molecule/virus/cell analyses.

Spectroscopy of the anharmonic cantilever oscillations in tapping-mode atomic-force microscopy

By spectroscopic analysis of the cantilever oscillation in tapping-mode atomic-force microscopy (TM–AFM), we demonstrate that the transition from an oscillatory state dominated by a net attractive force to the state dominated by repulsive interaction is accompanied by the enhanced generation of higher harmonics. The higher harmonics are a consequence of the nonlinear interaction and are amplified to significant amplitudes by the eigenmodes of the cantilever. The results show that in a quantitative description of TM–AFM higher eigenmode excitation must be considered to account for internal energy dissipation.