Cypris larvae of the barnacle Semibalanus balanoides leave proteinaceous footprints on surfaces during pre-settlement exploration. These footprints are considered to mediate temporary adhesion of cyprids to substrata and, as such, represent a crucial first step in the colonization of man-made surfaces by barnacles, a process known as biofouling. Interest in this system also stems from the potential for a synthetic reversible adhesion system, based on the strategy used by cyprids. Cyprid footprints were probed using atomic force microscopy (AFM) and nanomechanical data relating to interfacial adhesion forces were correlated with AFM tip chemistry. Commercial Si3N4-tips and chemically functionalized CH3-tips were chosen to mimic the interactions of cyprid footprints with hydrophilic and hydrophobic surfaces, respectively. Force-extension curves of protein bundles picked up by AFM tips exhibited a characteristic saw-tooth appearance for both types of tip, but demonstrated clear differences relating to pull-off force and pull-off length, based on tip chemistry. Additional (6nN) interfacial adhesion forces between -CH3 functionalized tips and footprints were assigned to hydrophobic interactions. Footprint proteins adhered with greater tenacity to the hydrophobic tip. This may suggest conformational change and denaturing of the protein which would facilitate hydrophobic interaction by enhancing contact forces between -CH3 functionalized tips and hydrophobic groups in the footprint molecule(s). Neither tip removed proteins from the -NH2 substratum suggesting that specific chemical interactions, rather than simple wetting phenomena, govern the adhesion of footprint proteins to that surface.