Phil M. Williams

In situ observation of streptavidin‐biotin binding on an immunoassay well surface using an atomic force microscope

Polystyrene microtitre wells are commonly used as supports for the enzyme‐linked immunosorbent assay (ELISA) method of biomolecular detection, which is employed in the routine diagnosis of a variety of medical conditions. We have used an atomic force microscope (AFM) to directly monitor specific molecular interactions between individual streptavidin and biotin molecules on such wells. This was achieved by functionalising an AFM probe with biotin and monitoring the adhesive forces between the probe and a streptavidin coated immunoassay well. The results demonstrate that the AFM may be employed as an analytical tool to study the interactions between biomolecules involved in immunoassay systems.

The use of a polymer film to estimate AFM probe profile

Abstract During our investigation of the surface morphology of various polyethylene glycol methacrylate copolymers we found that the rapid drying of a chloroform solution of a poly(methyl methacrylate- co -polyethylene glycol methacrylate) polymer resulted in the generation of a surface characterised by many sharp sided holes in a flat polymer matrix, which lead to extensive tip self imaging. By measuring the profiles of these holes (which are of variable size ranging from tens of nanometres to microns in depth), it is possible to estimate the gross tip profile over a wide vertical range in a totally non-destructive manner.

SDynamic Force Spectroscopy with the Atomic Force Microscope

By studying the effect of force on the dissociation kinetics of molecular interactions, hitherto hidden information about physics, chemistry, and biology is gained. Since the statement of the theory and the first demonstration of the experiment [2], we have seen developments in theory, experimental practice, and data analysis. Advances in theory have suggested the possibility of measuring more than dissociation rates over transition states and their displacements, such as the change in energy of the system at the transition state [3–5], the roughness of the dissociation landscape [6–8], and equilibrium phenomena [9]. Today, the instrumentation used to undertake DFS that is most prevalent in the literature is the atomic force microscope (AFM). The significant advances we have seen in both theory and experiment have sometimes taken place in isolation, and here I believe it is worth considering the application of DFS with current AFM technology. How accurately can we do DFS with an AFM? What exactly can we measure with the AFM, and what advances are needed?