Platelet activation plays a critical role in haemostasis and thrombosis. It is well-known that platelets generate contractile forces during activation. However, their mechanical material properties have rarely been investigated. Here, we use scanning ion conductance microscopy (SICM) to visualise morphological and mechanical properties of live human platelets at high spatial resolution. We found that their mean elastic modulus decreases during thrombin-induced activation by about a factor of two. We observed a similar softening of platelets during cytochalasin D-induced cytoskeleton depolymerisation. However, thrombin-induced temporal and spatial modulations of the elastic modulus were substantially different from cytochalasin D-mediated changes. We thereby provide new insights into the mechanics of haemostasis and establish SICM as a novel imaging platform for the ex vivo investigation of the mechanical properties of live platelets.

Nanotemplate directed DNA segmental thermal motion on molecular nanotemplates on graphite was directly observed and characterized using AFM in a liquid.

We developed resonance compensating chirp mode (RCCM), an atomic force microscopy (AFM) technique to measure the frequency dependence of the complex shear modulus of live cells over a large bandwidth (quasi-) continuously. RCCM works by applying a continuous frequency sweep (chirp) to the z-scanner and recording the resulting cantilever deflection at high speed. From this data, the frequency-resolved complex shear modulus is extracted. To reach a high maximum frequency, we iteratively shaped the chirp signal to compensate for scanner resonances. This allowed us to measure at frequencies five times higher than the resonant frequency of the scanner. Using a high-speed AFM with small cantilevers, we measured the complex shear modulus of live fibroblast cells in a continuous range between 5 Hz and 30 kHz. We found that the modulus and the loss tangent exhibit a power-law behavior throughout this frequency range. A short chirp duration of 200 ms allowed us to map live cells and generate spatially resolved images of the power-law parameters within minutes. These maps represent a unique combination of high spatial and frequency resolution, low measurement duration, and high maximum frequency.

Atomic force microscopy (AFM) of biomolecular processes at the single-molecule level can provide unique information for understanding molecular function. In AFM studies of biomolecular processes in solution, mica surfaces are predominantly used as substrates. However, owing to its high surface charge, mica may induce high local ionic strength in the vicinity of its surface, which may shift the equilibrium of studied biomolecular processes such as biopolymer adsorption or protein-DNA interaction. In the search for alternative substrates, we have investigated the behavior of adsorbed biomolecules, such as plasmid DNA and E. coli RNA polymerase σ subunit holoenzyme (RNAP), on highly oriented pyrolytic graphite (HOPG) surfaces modified with stearylamine and oligoglycine-hydrocarbon derivative (GM) monolayers using AFM in solution. We have demonstrated ionic-strength-dependent DNA mobility on GM HOPG and nativelike dimensions of RNAP molecules adsorbed on modified HOPG surfaces. We propose an approach to the real-time AFM investigation of transcription on stearylamine monolayers on graphite. We conclude that modified graphite allows us to study biomolecules and biomolecular processes on its surface at controlled ionic strength and may be used as a complement to mica in AFM investigations.

Current high‐speed atomic force microscopy (HS‐AFM) setups reach imaging speeds of several images per second but often have limited options for imaging live cells because of a small scan range, a lack of environmental control, or a missing combination with optical phase‐contrast or fluorescence microscopy. A HS‐AFM setup is therefore developed with a large scan range optimized for imaging live cells. The setup is equipped with temperature and CO2 control and is mounted on an inverted optical microscope providing high‐quality phase‐contrast and fluorescence microscopy. To demonstrate the capabilities of the setup, fast force mapping on live human platelets is performed. Further, HS‐AFM images and optical phase‐contrast and actin fluorescence images of live cancer cells are simultaneously recorded, and two state‐of‐the‐art AFM modes for imaging viscoelastic sample properties, force clamp force mapping and resonance compensating chirp mode, are compared. The setup is then applied to the investigation of viscoelastic material properties of cells in different cell cycle states. Using a melanoma cell line with a fluorescent cell cycle sensor, it is found that during the cell cycle not only cell volume and morphology, but also viscoelastic material properties significantly change, with increasing stiffness and decreasing fluidity from the G1 through the G1/S to the S/G2/M phases.