Here, we discuss the effects that the dynamics of the hydration layer and other variables, such as the tip radius, have on the availability of imaging regimes in dynamic AFM-including multifrequency AFM. Since small amplitudes are required for high-resolution imaging, we focus on these cases. It is possible to fully immerse a sharp tip under the hydration layer and image with amplitudes similar to or smaller than the height of the hydration layer, i.e., ~1 nm. When mica or HOPG surfaces are only cleaved, molecules adhere to their surfaces, and reaching a thermodynamically stable state for imaging might take hours. During these first hours, different possibilities for imaging emerge and change, implying that these conditions must be considered and reported when imaging.
Keywords: AFM; hydration; multifrequency; small amplitude; water.