Atomic Force Microscope to measure receptor - ligand interactions

Enormous progress has been made during the past two decades in our understanding of the cell surface and its components. Largely improved molecular biological techniques have led to the identification of numerous molecules expressed at the cell surface including ion channels, growth factor receptors, adhesion receptors, receptors of the immune system, as well as the ligands that bind to these various receptors. Despite this structural knowledge, we still have no clear insight in the receptor-ligand interactions at the molecular level. In particular the number of interactions required to induce signalling through receptors, the precise organisation of these receptor molecules at the cell surface, the forces between receptor-ligand pairs and most important the dynamics of receptor-ligand interactions. Until now such studies were hampered by the fact that current microscopical techniques that allow analysis of living cells have a maximal resolution of about 0.3 micron, depending on the light source used. Although electron microscopy provides much greater resolution, it does not allow the study of dynamic systems. Recent developments in scanning probe techniques including Atomic Force Microscopy now facilitate studies of biologicals (cells, organelles and isolated proteins) at high resolution in their natural environment. This allows for the first time real time analysis of biological systems at high nanometer resolution, such as single receptor-ligand interactions. In collaboration with the department of Technical Optics of the University of Twente (Prof. Dr. Greve) we have studied during the past 3 years the potential of AFM to investigate biologicals. Our results, and those emerging from recent literature, demonstrate that AFM has come of age and is extremely well suited not only to obtain high resolution topographical maps of biologicals, but also to perform actual measurement of single molecule interactions. Since now reliable commercial AFM's are available, allowing routine measurements by cell biologists without the need of a constant support of physical engineers we would like to introduce an AFM especially equipped for in situ studies by cell biologists within the Institute of Cellular Signalling to investigate biologicals at the molecular level.