Biological materials such as hexagonal packed intermediate (HPI) layers, DNA, tobacco mosaic virus and collagen deposited on various substrates with noncontact dynamic force microscopy under ambient conditions were investigated. This method is highly suited for the investigation of soft organic matter where a minimized interaction between tip and sample is needed for nondestructive and reliable operation. Hence, additional anchoring of the biological specimens was no longer found to be crucial. The vertical and lateral resolution limits of this gentle method were determined to be<0.1 nm and 1–2 nm, respectively, allowing very stable and high resolution results on all investigated systems. By taking approach curves and monitoring the dynamic properties of the cantilever (resonance frequency andQvalue) during the experiment, the interaction mechanism between tip and sample was found to be dominated by attractive van der Waals interaction and capillary forces. Furthermore, initial results from a HPI layer imaged with noncontact dynamic force microscopy in a water environment are presented.