Transport of fluids through channels and cavities of nano/mesoporous materials is of paramount importance in various fields of science and industry. The transport properties can be well derived from the adsorption− desorption behavior of fluids. Positron annihilation lifetime spectroscopy (PALS) allows probing adsorption−desorption of water from 2D mesopores of SBA-3. In situ study of the size of evolving water-free volumes during successive stages of adsorption and desorption is a sensitive way to elucidate the course of pore filling and emptying. The changes of positron annihilation parameters indicate that adsorption of water is mediated through the formation of isles on the surface of the pore walls, and these, in turn, develop into water plugs. Subsequently, these plugs grow and consecutively join together when the distance between them decreases to ca. 1 nm until the complete capillary condensation occurs. Akin to adsorption, desorption of water from the pores involves the formation of cavities capped with water plugs. The final stage of desorption shows the presence of water trapped in micropores in the pore walls. The linear dependence between the volume of water and the intensity of the water-related positronium component allows to estimate the amount of water in the system. The study highlights an approach to understand adsorption−desorption mechanism of liquids in mesopores by probing liquid-free volumes using ortho-positronium.