Biomedical subjects

Andrew D L Humphris

Publications and source records attributed to Andrew D L Humphris.

3 recordsLinked to original sources

PubMed2005

VideoAFM--a new tool for high speed surface analysis.

The VideoAFM provides a 1000 fold increase in image rate compared to conventional atomic force microscopes, giving nanometre resolution images of surfaces at a rate of 15 frames s(-1), which is approximately 1 million pixels s(-1). Images of high stiffness surfaces such as calibration grids are provided for the first time, and allow for a more rigorous examination of the meaning of the data obtained with the VideoAFM. Instrumental changes that could provide true topographic images are discussed. The advantages of a high speed scanning technique that is integrated within a conventional AFM are outlined. Particular emphasis is given to the capability to 'tile' images, and hence rapidly map large areas with nanometre resolution. It is found that the inherent increase in stability that comes from a high frame rate leads to the possibility of manually manipulating the sample while maintaining a sharp image, allowing real-time user interaction with the AFM. The possible application of the VideoAFM approach for the very rapid analysis of surface properties and, ultimately, surface chemistry is discussed and some possible routes are given.

Microscopy, Atomic Force↗

PubMed2005

Direct measurement of single-molecule visco-elasticity in atomic force microscope force-extension experiments.

Measuring the visco-elastic properties of biological macromolecules constitutes an important step towards the understanding of dynamic biological processes, such as cell adhesion, muscle function, or plant cell wall stability. Force spectroscopy techniques based on the atomic force microscope (AFM) are increasingly used to study the complex visco-elastic response of (bio-)molecules on a single-molecule level. These experiments either require that the AFM cantilever is actively oscillated or that the molecule is clamped at constant force to monitor thermal cantilever motion. Here we demonstrate that the visco-elasticity of single bio-molecules can readily be extracted from the Brownian cantilever motion during conventional force-extension measurements. It is shown that the characteristics of the cantilever determine the signal-to-noise (S/N) ratio and time resolution. Using a small cantilever, the visco-elastic properties of single dextran molecules were resolved with a time resolution of 8.3 ms. The presented approach can be directly applied to probe the dynamic response of complex bio-molecular systems or proteins in force-extension experiments.

Data Interpretation, Statistical↗

PubMed2004

Molecular force modulation spectroscopy revealing the dynamic response of single bacteriorhodopsins.

Recent advances in atomic force microscopy allowed globular and membrane proteins to be mechanically unfolded on a single-molecule level. Presented is an extension to the existing force spectroscopy experiments. While unfolding single bacteriorhodopsins from native purple membranes, small oscillation amplitudes (6-9 nm) were supplied to the vertical displacement of the cantilever at a frequency of 3 kHz. The phase and amplitude response of the cantilever-protein system was converted to reveal the elastic (conservative) and viscous (dissipative) contributions to the unfolding process. The elastic response (stiffness) of the extended parts of the protein were in the range of a few tens pN/nm and could be well described by the derivative of the wormlike chain model. Discrete events in the viscous response coincided with the unfolding of single secondary structure elements and were in the range of 1 microNs/m. In addition, these force modulation spectroscopy experiments revealed novel mechanical unfolding intermediates of bacteriorhodopsin. We found that kinks result in a loss of unfolding cooperativity in transmembrane helices. Reconstructing force-distance spectra by the integration of amplitude-distance spectra verified their position, offering a novel approach to detect intermediates during the forced unfolding of single proteins.

Bacteriorhodopsins↗