Quantitative Atomic Force Microscopy of Biochemical Binding Reactions
Atomic force microscopy has been used to generate atomic-resolution images of features and molecules present on atomically-flat surfaces such as mica or crystalline gold. In addition to vacuum or air-imaging instruments, newer instruments have the additional capability to image surfaces under liquids such as water, aqueous buffers and other solvents, and this has generated a great deal of interest in biomolecular characterization by AFM.
AFM experiments include contact-mode, in which a constant force is maintained between a cantilever tip and the sample surface by adjusting the height of the cantilever, and tapping-mode, where an oscillating cantilever is amplitude-damped by interactions with features on the sample surface. For delicate biological macromolecules, tapping-mode experiments can be used to generate three-dimensional representations of the structures of individual adsorbed molecules.
We are interested in using AFM to quantitate biomolecular binding. We have chosen a model reaction that occurs between Concanavalin A, a plant lectin, and polysaccharides such as dextran, a glucose polymer. Individual Con A tetramers are readily imaged on a mica surface under appropriate pH and ionic strength conditions, and the presence or absence of a polysaccharide attached to the Con A molecules can be determined by cross-sectional analysis of height images obtained using tapping-mode AFM.
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