Noncontact atomic force microscopy (AFM) using frequency modulation (FM) detection allows atomic resolution to be obtained in vacuum on a variety of insulating surfaces and molecular deposits. This technique has recently been extended to liquid environments, and, in addition to atomic scale contrast, FM-AFM in liquid allows measurement of ordered liquid layers above surfaces. The role of water and ions in biological processes is of great interest and in order to localize fluorescently tagged structures, such as proteins, optical microscopy combined with AFM provides an invaluable tool. Thus, to take advantage of the wealth of optical identification techniques available in biology, the AFM must be coupled to an optical microscope. Such systems are commercially available, but mechanical noise due to vibrations is a major concern compared with the compact, specialized instruments used to measure hydration structure to date. In this article the authors demonstrate, through both modeling and measurement, that hydration structure can be measured on such a commercial “bio-AFM,” despite the additional noise sources present in these instruments and that with the addition of a bandpass filter and amplifier it can be done “out-of-the-box” using only commercial electronics and tips. Thus, hydration structure measurements are accessible to virtually any laboratory with such a system.

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