Microscopy method fishes out individual proteins
could vastly improve the ability of atomic force microscopes to
visualise the chemical composition of a sample, follow variations
of the sample, as well as map its topographic structure.
They claim the advance could have significant implications for drug development by allowing scientists to monitor the effects of potential drugs on an ever-smaller scale.
"Atomic force microscopy (AFM) has a resolution down to an atomic level, but until now it has been blind to identifying specific chemical compositions," according to Stuart Lindsay, director of the Centre for Single Molecule Biophysics at the Biodesign Institute at Arizona State University and a lead researcher on the project.
The new technique allows an atomic force microscope to 'see', on a nanometer scale, the chemical composition of molecules.
Atomic force microscopes provide images on the nanometre scale by using a highly sensitive and tiny probe that is essentially pulled across a surface. By doing this, researchers can obtain topographical images down to a nanometer scale.
To use the AFM in its new mode, the researchers attached antibodies designed to bind to individual proteins to the tip of an AFM's probe using a tiny polymer thread. When the antibody binds to the target protein, it creates a variance in the microscope's reading compared to a reading with a bare tip, thus showing the presence of a protein or other specific material in the region being scanned.
Using a strand of polymer to connect the antibody to the tip provides flexibility, allowing the antibody to wiggle into position to better connect with the protein receptors. A magnetically excited cantilever makes the tip oscillate up and down to make the antibody disconnect and reconnect and keep the probe moving.
Lindsay describes the new capabilities of AFM by likening proteins to Lego bricks: conventional AFM can detect that there are bricks on the floor from their size and shape, but the improved version also tell what colour they are.
Giving an example of the limitations of AFM, he noted that in a sample of chromatin (nucleosomes) plus other proteins, DNA can be identified by its thread-like appearance, but the various protein components look similar, with an image size that depends only marginally on molecular weight.
A key capability of this technique, Lindsay said, is that it allows researchers to see how components of a cell react on a molecular scale when they experience biological processes, such as their response to a specific chemical or compound. In this mode, it could provide researchers with a molecular 'time-lapsed movie' of such reactions, which could lead to greater understanding of the chemical dynamics involved in how cells react to such stimuli.
Significant for drug discovery
"This development opens up the AFM as a research tool," Lindsay added. "The ability to identify the specific proteins on a membrane surface means you can take something very complex, like the surface of a human cell with all of the types of different receptors on it and ask questions about the local chemistry, like what is binding at those sites. That will provide the fundamental knowledge you need to develop new drugs."
Lindsay said that the new process gives AFM chemical sensitivity at the nanometer scale, in much the way colored or fluorescent dyes gave conventional microscopes optical sensitivity at the micron scale.
At the time of writing the full article is available on the Proceedings of the National Academy of Sciences' website.
