Investigating the role of proteoglycans associated to cell membranes is fundamental to comprehend biochemical process that occurs at the level of membrane surfaces. In this paper, we exploit syndecan-4, a heparan sulfate proteoglycan obtained from cell cultures, in lipid Langmuir monolayers at the air-water interface. The monolayer served as a model for half a membrane, and the molecular interactions involved could be evaluated with tensiometry and vibrational spectroscopy techniques. Polarization-modulation infrared reflection-absorption spectroscopy (PM-IRRAS) employed in a constant surface pressure regime showed that the main chemical groups for syndecan-4 were present at the air-water interface. Subsequent monolayer decompression and compression showed surface pressure-area isotherms with a large expansion for the lipid monolayers interacting with the cell culture reported to over-express syndecan-4, which was also an indication that the proteoglycan was inserted in the lipid monolayer. The introduction of biological molecules with affinity for syndecam-4, such as growth factors, which present a key role in biochemical process of cell signaling, changed the surface properties of the hybrid film, leading to a model, by which the growth factor binds to the sulfate groups present in the heparan sulfate chains. The polypeptide moiety of syndecan-4 responds to this interaction changing its conformation, which leads to lipid film relaxation and further monolayer condensation.