Atomic force microscopy has been employed for the in situ investigation of the molecular hydration of thiolated dextran monolayers within a liquid environment. The studies have demonstrated the ability to measure corresponding changes in both monolayer morphology and elasticity due to the hydration state of dextran. Imaging in water allows the visualization of macromolecular swelling, matched by an increase in surface elasticity monitored via the analysis of force-distance curves. Subsequent imaging in a propanol environment leads to dehydration effects observed through a relaxation of swelling and a decrease in surface elasticity.